Prmt5 inhibitors

ABSTRACT

The present invention provides a compound of Formula (I) or the pharmaceutically acceptable salts thereof, which are PRMT5 inhibitors.

BACKGROUND OF THE INVENTION

PRMT5 (aka JBP1, SKB1, 1BP72, SKB1his and HRMTIL5) is a Type II argininemethyltransferase, and was first identified in a two-hybrid search forproteins interacting with the Janus tyrosine kinase (Jak2) (Pollack etal., 1999). PRMT5 plays a significant role in control and modulation ofgene transcription. Inter alia, PRMT5 is known to methylate histone H3at Arg-8 (a site distinct from that methylated by PRMT4) and histone H4at Arg-3 (the same site methylated by PRMT1) as part of a complex withhuman SWI/SNF chromatin remodelling components BRG1 and BRM.

PRMT5 is involved in the methylation and functional modulation of thetumor suppressor protein p53. (See Berger, 2008; Durant et al., 2009;Jansson et al., 2008; Scoumanne et al., 2009). Most of the physiologicalfunctions of p53 are attributable to its role as a transcriptionalactivator, responding to agents that damage DNA. p53 status is wild typein approximately half of human cancer cases. These include 94% incervix, 87% in blood malignancies, 85% in bones and endocrine glands,and 75% of primary breast cancer. Restoration of p53 in cancer cellsharbouring wild type p53, by way of inhibiting mechanisms that suppressits function leads to growth arrest and apoptosis, and is regarded as apotentially effective means of tumor suppression.

The role of PRMT5 and p53 arginine methylation on cell cycle regulationand DNA damage response have been explored by both Jansson et al. andScoumanne et al. (Jansson et al., 2008; Scoumanne et al., 2009).Although some differences are evident between the results from the twogroups concerning cell cycle regulation in unperturbed cells (which maybe ascribed to cell type specific effects and/or the actual nature ofthe experimental arrangements), both groups report similar results withrespect to the DNA damage response.

In response to DNA damage caused by a variety of agents, includingdoxorubicin, camptothecin and UV light, and also in response totreatment with Nutlin-3, knockdown of PRMT5 results in an increase insub-G1 population and concomitant reduction in G1 cells and, in thepresence of p53, a significant increase in apoptosis. Knockdown of PRMT5also resulted in a reduced level of p21, a key p53 target gene thatregulates cell cycle arrest during the p53 response and MDM2, a p53 E3ubiquitin ligase, but not PUMA, NOXA, AlP1 & APAF1, p53 target geneslinked to apoptosis.

Knockdown of PRMT5 (but not PRMT1 or CARM1/PRMT4) results in decreasedp53 stabilisation, decreased basal p53 levels, decreased p53oligomerisation, and also decreased expression of elF4E a majorcomponent of translational machinery involved in ribosome binding tomRNA. Indeed, elF4E is a potent oncogene, which has been shown topromote malignant transformation in vitro and human cancer formation.

Knockdown of PRMT5 would be expected to lead to a reduction in the levelof arginine methylated p53. Consistent with arginine methylation statusof p53 influencing the p53 response (reduced arginine methylationbiasing the response to proapoptotic), Jannson et al. showed that a p53mutant in which each of the three critical arginine residues weresubstituted with lysine (p53KKK) retained the ability to induceapoptosis but its cell cycle arrest activity was significantlycompromised.

Moreover, pS3KKK also has a significantly reduced ability to inducetranscription of p21, by contrast with APAF1. The promoter bindingspecificity of wild-type p53 to key target genes is also significantlyaffected by arginine methylating status: Knockdown of PRMT5 results indecreased p53 binding to the promoter regions of the p21 and(intriguingly) PUMA genes, but does not affect p53 binding to thepromoter regions of NOXA or APAF1.

PRMT5 is aberrantly expressed in around half of human cancer cases,further linking this mechanism to cancers. PRMT5 overexpression has beenobserved in patient tissue samples and cell lines of Prostate cancer (Guet al., 2012), Lung cancer (Zhongping et al., 2012), Melanoma cancer(Nicholas et al., 2012), Breast cancer (Powers et al., 2011), Colorectalcancer (Cho et al., 2012), Gastric cancer (Kim et al., 2005), Esophagusand Lung carcinoma (Aggarwal et al., 2010) and B-Cell lymphomas andleukemia (Wang, 2008). Moreover, elevated expression of PRMT5 inMelanoma, Breast and Colorectal cancers has been demonstrated tocorrelate with a poor prognosis.

Lymphoid malignancies including chronic lymphcytic leukemia (CLL) areassociated with over-expression of PRMT5. PRMT5 is over-expressed (atthe protein level) in the nucleus and cytosol in a number of patientderived Burkitt's lymphoma; mantle cell lymphoma (MCL); in vitroEBV-transformed lymphoma; leukaemia cell lines; and B-CLL cell lines,relative to normal CD19+B lymphocytes (Pal et al., 2007; Wang et al.,2008). Intriguingly, despite elevated levels of PRMT5 protein in thesetumor cells, the levels of PRMT5 mRNA are reduced (by a factor of 2-5).Translation of PRMT5 mRNA is, however, enhanced in lymphoma cells,resulting in increased levels of PRMT5 (Pal et al., 2007; Wang et al.,2008).

In addition to genomic changes, CLL, like almost all cancers, hasaberrant epigenetic abnormalities characterised by globalhypomethylation and hot-spots of repressive hypermethylation ofpromoters including tumor suppressor genes. While the role ofepigenetics in the origin and progression of CLL remains unclear,epigenetic changes appear to occur early in the disease and specificpatterns of DNA methylation are associated with worse prognosis (Chen etal., 2009; Kanduri et al., 2010). Global symmetric methylation ofhistones H3R8 and H4R3 is increased in transformed lymphoid cell linesand MCL clinical samples (Pal et al., 2007), correlating with theoverexpression of PRMT5 observed in a wide variety of lymphoid cancercell lines and MCL clinical samples.

PRMT5 is therefore a target for the identification of novel cancertherapeutics.

Hemoglobin is a major protein in red blood cells and is essential forthe transport of oxygen from the lungs to the tissues. In adult humans,the most common hemoglobin type is a tetramer called hemoglobin A,consisting of two α and two β subunits. In human infants, the hemoglobinmolecule is made up of two α and two γ chains. The gamma chains aregradually replaced by subunits as the infant grows. The developmentalswitch in human B-like globin gene subtype from foetal (γ) to adult (β)that begins at birth heralds the onset of the hemoglobinopathiesβ-thalassemia and sickle cell disease (SCD). In β-thalassemia the adultchains are not produced. In SCD a point mutation in the coding sequencein the B globin gene leads to the production of a protein with alteredpolymerisation properties. The observation that increased adult γ-globingene expression (in the setting of hereditary persistence of foetalhemoglobin (HPFH) mutations) significantly ameliorates the clinicalseverity of β-thalassemia and SCD has prompted the search fortherapeutic strategies to reverse γ-globin gene silencing. To date, thishas been achieved through pharmacological induction, using compoundsthat broadly influence epigenetic modifications, including DNAmethylation and histone deacetylation. The development of more targetedtherapies is dependent on the identification of the molecular mechanismsunderpinning foetal globin gene silencing. These mechanisms haveremained elusive, despite exhaustive study of the HPFH mutations, andconsiderable progress in many other aspects of globin gene regulation.

PRMT5 plays a critical role in triggering coordinated repressiveepigenetic events that initiate with dimethylation of histone H4Arginine 3 (H4R3me2s), and culminate in DNA methylation andtranscriptional silencing of the γ-genes (Rank et al., 2010). Integralto the synchronous establishment of the repressive markers is theassembly of a PRMT5-dependent complex containing the DNAmethyltransferase DNMT3A, and other repressor proteins (Rank et al.,2010). DNMT3A is directly recruited to bind to the PRMT5-inducedH4R3me2s mark, and loss of this mark through shRNA-mediated knock-downof PRMT5, or enforced expression of a mutant form of PRMT5 lackingmethyltransferase activity leads to marked upregulation of γ-geneexpression, and complete abrogation of DNA methylation at theγ-promoter. Treatment of human erythroid progenitors with non-specificmethyltransferase inhibitors (Adox and MTA) also resulted inupregulation of γ-gene expression (He Y, 2013). Inhibitors of PRMT5 thushave potential as therapeutics for hemoglobinopathies such asβ-thalassemia and Sickle Cell Disease (SCD).

The present inventors have developed particular tetrahydroisoquinolinesthat inhibit the activity of PRMT5 and therefore may be of use intreating conditions ameliorated by the inhibition of the activity ofPRMT5.

SUMMARY OF THE INVENTION

Compounds of formula I

or the pharmaceutically acceptable salts, esters, and prodrugs thereof,which are PRMT5 inhibitors. Also provided are pharmaceuticalcompositions comprising compounds of Formula I, and methods of usingthese compounds to treat cancer.

DETAILED DESCRIPTION OF THE INVENTION

Compounds of formula I

or pharmaceutically acceptable salts thereof, whereinR¹ is hydrogen or C₁₋₄ alkyl, andR² is hydrogen or C₁₋₄ alkyl, orR¹ and R², together with the carbon atom to which they are attached,form C₃₋₈ cycloalkyl;

Z is

halogen,

C(O)R³,

a 5-membered unsaturated heterocycle, or

an 8-membered bridged bicyclic saturated heterocycle; and

R³ is

an 8-membered bridged bicyclic saturated heterocycle,

an 8-membered bridged bicyclic heterocycle, or

a 9-membered bridged bicyclic saturated heterocycle; and

R⁶, R⁷, R⁸, and R⁹, are independently selected from hydrogen, halogenand C₁₋₄ alkyl.

In an embodiment of the invention is a compound of Formula I,

or a pharmaceutically acceptable salt thereof, whereinR¹ is hydrogen or C₁₋₄ alkyl, andR² is hydrogen or C₁₋₄ alkyl, orR and R², together with the carbon atom to which they are attached, formC₃₋₈ cycloalkyl;

Z is

-   -   halogen,    -   C(O)R³,    -   a 5-membered unsaturated heterocycle containing 3 N atoms,        wherein the heterocycle is unsubstituted or substituted with CH₃        or cyclopropyl, or    -   an 8-membered bridged bicyclic saturated heterocycle containing        1 N atom, unsubstituted or substituted with ═O; and

R³ is

-   -   an 8-membered bridged bicyclic saturated unsubstituted or        substituted heterocycle containing 1N atom and 1 O atom,    -   an 8-membered bridged bicyclic heterocycle containing 1 N atom,        wherein the heterocycle is unsubstituted or substituted with OH        or OCH₃, or    -   a 9-membered bridged bicyclic saturated unsubstituted or        substituted heterocycle containing 1 N atom and 2 O atoms,    -   wherein the substituted heterocycle is substituted with one,        two, or three moieties independently selected from halogen; and        R⁶, R⁷, R⁸, and R⁹, are independently selected from hydrogen,        halogen and C₁₋₄ alkyl.

In an embodiment of the invention,

R³ is

-   -   an 8-membered bridged bicyclic saturated unsubstituted        heterocycle containing 1N atom and 1 O atom,    -   an 8-membered bridged bicyclic heterocycle containing 1 N atom,        wherein the heterocycle is unsubstituted or substituted with OH        or OCH₃, or    -   a 9-membered bridged bicyclic saturated heterocycle containing 1        N atom and 2 O atoms; and        R⁶, R⁷, R⁸, and R⁹, are hydrogen.

In an embodiment of the invention, R¹ is hydrogen or CH₃ and R² ishydrogen, CH₃, or CH₂CH₃ or R¹ and R², together with the carbon atom towhich they are attached, form cyclobutyl.

In an embodiment of the invention, R³ is

In an embodiment of the invention, Z is Br,

In an embodiment of the invention, the compounds is

The term “composition” is intended to encompass a product comprising thespecified ingredients in the specified amounts, as well as any productwhich results, directly or indirectly, from combination of the specifiedingredients in the specified amounts. The term “anti-cancer agent” meansa drug (medicament or pharmaceutically active ingredient) for treatingcancer. The term “antineoplastic agent” means a drug (medicament orpharmaceutically active ingredient) for treating cancer (i.e., achemotherapeutic agent). The term “at least one” means one or more thanone. The meaning of “at least one” with reference to the number ofcompounds of the invention is independent of the meaning with referenceto the number of chemotherapeutic agents. The term “chemotherapeuticagent” means a drug (medicament or pharmaceutically active ingredient)for treating cancer (i.e., an antineoplastic agent). The term “compound”with reference to the antineoplastic agents, includes the agents thatare antibodies. The term “consecutively” means one following the other.The term “effective amount” means a “therapeutically effective amount”.The term “therapeutically effective amount” means that amount of activecompound or pharmaceutical agent that elicits the biological ormedicinal response in a tissue, system, animal or human that is beingsought by a researcher, veterinarian, medical doctor or other clinician.Thus, for example, in the methods of treating cancer described herein“effective amount” (or “therapeutically effective amount”) means, theamount of the compound (or drug), or radiation, that results in: (a) thereduction, alleviation or disappearance of one or more symptoms causedby the cancer, (b) the reduction of tumor size, (c) the elimination ofthe tumor, and/or (d) long-term disease stabilization (growth arrest) ofthe tumor. Also, for example, an effective amount, or a therapeuticallyeffective amount of the PRMT5 inhibitor (i.e., a compound of theinvention) is that amount which results in the reduction in PRMT5activity. The term “treating cancer” or “treatment of cancer” refers toadministration to a mammal afflicted with a cancerous condition andrefers to an effect that alleviates the cancerous condition by killingthe cancerous cells, and also refers to an effect that results in theinhibition of growth and/or metastasis of the cancer.

The invention also provides a pharmaceutical composition comprising aneffective amount of at least one compound of Formula I and apharmaceutically acceptable carrier. The invention also provides apharmaceutical composition comprising an effective amount of at leastone compound of Formula I and an effective amount of at least one otherpharmaceutically active ingredient (such as, for example, achemotherapeutic agent), and a pharmaceutically acceptable carrier.

The invention also provides a method of inhibiting PRMT5 in a patient inneed of such treatment comprising administering to said patient aneffective amount of at least one compound of Formula I. The inventionalso provides a method for treating cancer in a patient in need of suchtreatment, said method comprising administering to said patient aneffective amount of at least one compound of Formula I. The inventionalso provides a method for treating cancer in a patient in need of suchtreatment, said method comprising administering to said patient aneffective amount of at least one compound of Formula I, in combinationwith an effective amount of at least one chemotherapeutic agent. Themethods of the invention include the administration of a pharmaceuticalcomposition comprising at least one compound of the invention and apharmaceutically acceptable carrier. The invention also provides any ofthe above methods of treating cancer wherein the cancer is colorectal.The invention also provides any of the above methods of treating cancerwherein the cancer is melanoma. The methods of treating cancersdescribed herein can optionally include the administration of aneffective amount of radiation (i.e., the methods of treating cancersdescribed herein optionally include the administration of radiationtherapy).

The methods of treating cancer described herein include methods oftreating cancer that comprise administering a therapeutically effectiveamount of a compound of the instant invention in combination withradiation therapy and/or in combination with a second compound selectedfrom: an estrogen receptor modulator, an androgen receptor modulator, aretinoid receptor modulator, a cytotoxicytostatic agent, anantiproliferative agent, a prenyl-protein transferase inhibitor, anHMG-CoA reductase inhibitor, an HIV protease inhibitor, a reversetranscriptase inhibitor, an angiogenesis inhibitor, PPAR-γ agonists,PPAR-δ agonists, an inhibitor of inherent multidrug resistance, ananti-emetic agent, an agent useful in the treatment of anemia, an agentuseful in the treatment of neutropenia, an immunologic-enhancing drug,an inhibitor of cell proliferation and survival signaling, abisphosphonate, an aromatase inhibitor, an siRNA therapeutic,γ-secretase and/or NOTCH inhibitors, agents that interfere with receptortyrosine kinases (RTKs), an agent that interferes with a cell cyclecheckpoint, and any of the therapeutic agents listed herein.

In any of the methods of treating cancer described herein, unless statedotherwise, the methods can optionally include the administration of aneffective amount of radiation therapy. For radiation therapy,γ-radiation is preferred.

Thus, another example of the invention is directed to a method oftreating cancer in a patient in need of such treatment, said methodcomprising administering an effective amount of a compound of Formula I.Another example of the invention is directed to a method of treatingcancer in a patient in need of such treatment, said method comprisingadministering to said patient an effective amount of a compound ofFormula I, and an effective amount of at least one chemotherapeuticagent.

Methods for the safe and effective administration of most of thesechemotherapeutic agents are known to those skilled in the art. Inaddition, their administration is described in the standard literature.For example, the administration of many of the chemotherapeutic agentsis described in the “Physicians' Desk Reference” (PDR), e.g., 1996edition (Medical Economics Company, Montvale, N.J. 07645-1742, USA), thePhysicians' Desk Reference, 56th Edition, 2002 (published by MedicalEconomics company, Inc. Montvale, N.J. 07645-1742), the Physicians' DeskReference, 57^(th) Edition, 2003 (published by Thompson PDR, Montvale,N.J. 07645-1742), the Physicians' Desk Reference, 60^(th) Edition, 2006(published by Thompson PDR, Montvale, N.J. 07645-1742), and thePhysicians' Desk Reference, 64^(th) Edition, 2010 (published by PDRNetwork, LLC at Montvale, N.J. 07645-1725); the disclosures of which areincorporated herein by reference thereto.

If the patient is responding, or is stable, after completion of thetherapy cycle, the therapy cycle can be repeated according to thejudgment of the skilled clinician. Upon completion of the therapycycles, the patient can be continued on the compounds of the inventionat the same dose that was administered in the treatment protocol. Thismaintenance dose can be continued until the patient progresses or can nolonger tolerate the dose (in which case the dose can be reduced and thepatient can be continued on the reduced dose).

Those skilled in the art will recognize that the actual dosages andprotocols for administration employed in the methods of the inventionmay be varied according to the judgment of the skilled clinician. Theactual dosage employed may be varied depending upon the requirements ofthe patient and the severity of the condition being treated.Determination of the proper dosage for a particular situation is withinthe skill of the art. A determination to vary the dosages and protocolsfor administration may be made after the skilled clinician takes intoaccount such factors as the patient's age, condition and size, as wellas the severity of the cancer being treated and the response of thepatient to the treatment.

The amount and frequency of administration of the compound of formula(1) and the chemotherapeutic agents will be regulated according to thejudgment of the attending clinician (physician) considering such factorsas age, condition and size of the patient as well as severity of thecancer being treated.

The chemotherapeutic agent can be administered according to therapeuticprotocols well known in the art. It will be apparent to those skilled inthe art that the administration of the chemotherapeutic agent can bevaried depending on the cancer being treated and the known effects ofthe chemotherapeutic agent on that disease. Also, in accordance with theknowledge of the skilled clinician, the therapeutic protocols (e.g.,dosage amounts and times of administration) can be varied in view of theobserved effects of the administered therapeutic agents on the patient,and in view of the observed responses of the cancer to the administeredtherapeutic agents.

The initial administration can be made according to establishedprotocols known in the art, and then, based upon the observed effects,the dosage, modes of administration and times of administration can bemodified by the skilled clinician.

The particular choice of chemotherapeutic agent will depend upon thediagnosis of the attending physicians and their judgement of thecondition of the patient and the appropriate treatment protocol.

The determination of the order of administration, and the number ofrepetitions of administration of the chemotherapeutic agent during atreatment protocol, is well within the knowledge of the skilledphysician after evaluation of the cancer being treated and the conditionof the patient.

Thus, in accordance with experience and knowledge, the practicingphysician can modify each protocol for the administration of achemotherapeutic agent according to the individual patient's needs, asthe treatment proceeds. All such modifications are within the scope ofthe present invention.

The attending clinician, in judging whether treatment is effective atthe dosage administered, will consider the general well-being of thepatient as well as more definite signs such as relief of cancer-relatedsymptoms (e.g., pain), inhibition of tumor growth, actual shrinkage ofthe tumor, or inhibition of metastasis. Size of the tumor can bemeasured by standard methods such as radiological studies, e.g., CAT orMRI scan, and successive measurements can be used to judge whether ornot growth of the tumor has been retarded or even reversed. Relief ofdisease-related symptoms such as pain, and improvement in overallcondition can also be used to help judge effectiveness of treatment.

The compounds, compositions and methods provided herein are useful forthe treatment of cancer. Cancers that may be treated by the compounds,compositions and methods disclosed herein include, but are not limitedto: (1) Cardiac: sarcoma (angiosarcoma, fibrosarcoma, rhabdomyosarcoma,liposarcoma), myxoma, rhabdomyoma, fibroma, lipoma and teratoma; (2)Lung: bronchogenic carcinoma (squamous cell, undifferentiated smallcell, undifferentiated large cell, adenocarcinoma), alveolar(bronchiolar) carcinoma, bronchial adenoma, sarcoma, lymphoma,chondromatous hamartoma, mesothelioma, non-small cell; (3)Gastrointestinal: esophagus (squamous cell carcinoma, adenocarcinoma,leiomyosarcoma, lymphoma), stomach (carcinoma, lymphoma,leiomyosarcoma), pancreas (ductal adenocarcinoma, insulinoma,glucagonoma, gastrinoma, carcinoid tumors, vipoma), small bowel(adenocarcinoma, lymphoma, carcinoid tumors, Karposi's sarcoma,leiomyoma, hemangioma, lipoma, neurofibroma, fibroma), large bowel(adenocarcinoma, tubular adenoma, villous adenoma, hamartoma,leiomyoma), colon, colorectal, rectal; (4) Genitourinary tract: kidney(adenocarcinoma, Wilm's tumor [nephroblastoma], lymphoma, leukemia),bladder and urethra (squamous cell carcinoma, transitional cellcarcinoma, adenocarcinoma), prostate (adenocarcinoma, sarcoma), testis(seminoma, teratoma, embryonal carcinoma, teratocarcinoma,choriocarcinoma, sarcoma, interstitial cell carcinoma, fibroma,fibroadenoma, adenomatoid tumors, lipoma); (5) Liver: hepatoma(hepatocellular carcinoma), cholangiocarcinoma, hepatoblastoma,angiosarcoma, hepatocellular adenoma, hemangioma; (6) Bone: osteogenicsarcoma (osteosarcoma), fibrosarcoma, malignant fibrous histiocytoma,chondrosarcoma, Ewing's sarcoma, malignant lymphoma (reticulum cellsarcoma), multiple myeloma, malignant giant cell tumor chordoma,osteochronfroma (osteocartilaginous exostoses), benign chondroma,chondroblastoma, chondromyxofibroma, osteoid osteoma and giant celltumors; (7) Nervous system: skull (osteoma, hemangioma, granuloma,xanthoma, osteitis deformans), meninges (meningioma, meningiosarcoma,gliomatosis), brain (astrocytoma, medulloblastoma, glioma, ependymoma,germinoma [pinealoma], glioblastoma multiform, oligodendroglioma,schwannoma, retinoblastoma, congenital tumors), spinal cordneurofibroma, meningioma, glioma, sarcoma); (8) Gynecological: uterus(endometrial carcinoma), cervix (cervical carcinoma, pre-tumor cervicaldysplasia), ovaries (ovarian carcinoma [serous cystadenocarcinoma,mucinous cystadenocarcinoma, unclassified carcinoma], granulosa-thecalcell tumors, Sertoli-Leydig cell tumors, dysgerminoma, malignantteratoma), vulva (squamous cell carcinoma, intraepithelial carcinoma,adenocarcinoma, fibrosarcoma, melanoma), vagina (clear cell carcinoma,squamous cell carcinoma, botryoid sarcoma (embryonal rhabdomyosarcoma),fallopian tubes (carcinoma), breast; (9) Hematologic: blood (myeloidleukemia [acute and chronic], acute lymphoblastic leukemia, chroniclymphocytic leukemia, chronic myelomonocytic (CMML), myeloproliferativediseases, multiple myeloma, myelodysplastic syndrome), Hodgkin'sdisease, non-Hodgkin's lymphoma [malignant lymphoma]; (10) Skin:malignant melanoma, basal cell carcinoma, squamous cell carcinoma,Karposi's sarcoma, moles dysplastic nevi, lipoma, angioma,dermatofibroma, keloids, psoriasis; and (11) Adrenal glands:neuroblastoma. Examples of cancer that may be treated by the compounds,compositions and methods of the invention include thyroid cancer,anaplastic thyroid carcinoma, epidermal cancer, head and neck cancer(e.g., squamous cell cancer of the head and neck), sarcoma,tetracarcinoma, hepatoma and multiple myeloma. Thus, the term “cancerouscell” as provided herein, includes a cell afflicted by any one of theabove-identified conditions.

In the treatment of breast cancer (e.g., postmenopausal andpremenopausal breast cancer, e.g., hormone-dependent breast cancer) thecompound of formula (1) may be used with an effective amount of at leastone antihormonal agent selected from the group consisting of: (a)aromatase inhibitors, (b) antiestrogens, and (c) LHRH analogues; andoptionally an effective amount of at least one chemotherapeutic agent.Examples of aromatase inhibitors include but are not limited to:Anastrozole (e.g., Arimidex), Letrozole (e.g., Femara), Exemestane(Aromasin), Fadrozole and Formestane (e.g., Lentaron). Examples ofantiestrogens include but are not limited to: Tamoxifen (e.g.,Nolvadex), Fulvestrant (e.g., Faslodex), Raloxifene (e.g., Evista), andAcolbifene. Examples of LHRH analogues include but are not limited to:Goserelin (e.g., Zoladex) and Leuprolide (e.g., Leuprolide Acetate, suchas Lupron or Lupron Depot). Examples of chemotherapeutic agents includebut are not limited to: Trastuzumab (e.g., Herceptin), Gefitinib (e.g.,Iressa), Erlotinib (e.g., Erlotinib HCl, such as Tarceva), Bevacizumab(e.g., Avastin), Cetuximab (e.g., Erbitux), and Bortezomib (e.g.,Velcade).

In one example of the invention the cancer treated is colo-rectal cancer(such as, for example, colon adenocarcinoma and colon adenoma). Thus,another example of the invention is directed to a method of treatingcolo-rectal cancer in a patient in need of such treatment, said methodcomprising administering an effective of a compound of Formula I, or apharmaceutically acceptable salt thereof, to said patient. Anotherexample of the invention is directed to a method of treating colo-rectalcancer in a patient in need of such treatment, said method comprisingadministering to said patient an effective amount of a compound ofFormula I, or a pharmaceutically acceptable salt thereof, and aneffective amount of at least one chemotherapeutic agent.

In one example of the invention the cancer treated is melanoma. Thus,another example of the invention is directed to a method of treatingmelanoma in a patient in need of such treatment, said method comprisingadministering an effective amount of a compound of Formula I, or apharmaceutically acceptable salt thereof, to said patient. Anotherexample of the invention is directed to a method of treating melanoma ina patient in need of such treatment, said method comprisingadministering to said patient an effective amount of a compound ofFormula I, or a pharmaceutically acceptable salt thereof, and aneffective amount of at least one chemotherapeutic agent.

The compounds of the invention are also useful in preparing a medicamentthat is useful in treating cancer.

The instant compounds are also useful in combination with therapeutic,chemotherapeutic and anti-cancer agents. Combinations of the presentlydisclosed compounds with therapeutic, chemotherapeutic and anti-canceragents are within the scope of the invention. Examples of such agentscan be found in Cancer Principles and Practice of Oncology by VT. Devitaand S. Hellman (editors), 6^(th) edition (Feb. 15, 2001), LippincottWilliams & Wilkins Publishers. A person of ordinary skill in the artwould be able to discern which combinations of agents would be usefulbased on the particular characteristics of the drugs and the cancerinvolved. Such agents include the following: estrogen receptormodulators, programmed cell death protein 1 (PD-1) inhibitors,programmed death-ligand 1 (PD-L1) inhibitors, androgen receptormodulators, retinoid receptor modulators, cytotoxic/cytostatic agents,antiproliferative agents, prenyl-protein transferase inhibitors, HMG-CoAreductase inhibitors and other angiogenesis inhibitors, HIV proteaseinhibitors, reverse transcriptase inhibitors, inhibitors of cellproliferation and survival signaling, bisphosphonates, aromataseinhibitors, siRNA therapeutics, γ-secretase inhibitors, agents thatinterfere with receptor tyrosine kinases (RTKs) and agents thatinterfere with cell cycle checkpoints. The instant compounds areparticularly useful when co-administered with radiation therapy.

“Estrogen receptor modulators” refers to compounds that interfere withor inhibit the binding of estrogen to the receptor, regardless ofmechanism. Examples of estrogen receptor modulators include, but are notlimited to, tamoxifen, raloxifene, idoxifene, LY353381, LY117081,toremifene, fulvestrant,4-[7-(2,2-dimethyl-1-oxopropoxy-4-methyl-2-[4-[2-(1-piperidinyl)ethoxy]phenyl]-2H-1-benzopyran-3-yl]-phenyl-2,2-dimethylpropanoate,4,4′-dihydroxybenzophenone-2,4-dinitrophenyl-hydrazone, and SH646.

PD-1 inhibitors include pembrolizumab (lambrolizumab), nivolumab andMPDL3280A. PDL-inhibitors include atezolizumab, avelumab, anddurvalumab.

“Androgen receptor modulators” refers to compounds which interfere orinhibit the binding of androgens to the receptor, regardless ofmechanism. Examples of androgen receptor modulators include finasterideand other 5α-reductase inhibitors, nilutamide, flutamide, bicalutamide,liarozole, and abiraterone acetate.

“Retinoid receptor modulators” refers to compounds which interfere orinhibit the binding of retinoids to the receptor, regardless ofmechanism. Examples of such retinoid receptor modulators includebexarotene, tretinoin, 13-cis-retinoic acid, 9-cis-retinoic acid,α-difluoromethylornithine, ILX23-7553, trans-N-(4′-hydroxyphenyl)retinamide, and N-4-carboxyphenyl retinamide.

“Cytotoxic/cytostatic agents” refer to compounds which cause cell deathor inhibit cell proliferation primarily by interfering directly with thecell's functioning or inhibit or interfere with cell myosis, includingalkylating agents, tumor necrosis factors, intercalators, hypoxiaactivatable compounds, microtubule inhibitors/microtubule-stabilizingagents, inhibitors of mitotic kinesins, histone deacetylase inhibitors,inhibitors of kinases involved in mitotic progression, inhibitors ofkinases involved in growth factor and cytokine signal transductionpathways, antimetabolites, biological response modifiers,hormonal/anti-hormonal therapeutic agents, haematopoietic growthfactors, monoclonal antibody targeted therapeutic agents, topoisomeraseinhibitors, proteosome inhibitors, ubiquitin ligase inhibitors, andaurora kinase inhibitors.

Examples of cytotoxic/cytostatic agents include, but are not limited to,sertenef, cachectin, ifosfamide, tasonermin, lonidamine, carboplatin,altretamine, prednimustine, dibromodulcitol, ranimustine, fotemustine,nedaplatin, oxaliplatin, temozolomide, heptaplatin, estramustine,improsulfan tosilate, trofosfamide, nimustine, dibrospidium chloride,pumitepa, lobaplatin, satraplatin, profiromycin, cisplatin, irofulven,dexifosfamide, cis-aminedichloro(2-methyl-pyridine)platinum,benzylguanine, glufosfamide, GPX100, (trans, trans,trans)-bis-mu-(hexane-1,6-diamine)-mu-[diamine-platinum(II)]bis[diamine(chloro)platinum(II)]tetrachloride, diarizidinylspermine, arsenic trioxide,1-(11-dodecylamino-10-hydroxyundecyl)-3,7-dimethylxanthine, zorubicin,idarubicin, daunorubicin, bisantrene, mitoxantrone, pirarubicin,pinafide, valrubicin, amrubicin, antineoplaston,3′-deamino-3′-morpholino-13-deoxo-10-hydroxycarminomycin, annamycin,galarubicin, elinafide, MEN10755,4-demethoxy-3-deamino-3-aziridinyl-4-methylsulphonyl-daunorubicin (seeWO 00/50032), Raf kinase inhibitors (such as Bay43-9006) and mTORinhibitors (such as Wyeth's CCI-779).

An example of a hypoxia activatable compound is tirapazamine.

Examples of proteosome inhibitors include but are not limited tolactacystin and MLN-341 (Velcade).

Examples of microtubule inhibitors/microtubule-stabilising agentsinclude paclitaxel, vindesine sulfate,3′,4′-didehydro-4′-deoxy-8′-norvincaleukoblastine, docetaxol, rhizoxin,dolastatin, mivobulin isethionate, auristatin, cemadotin, RPR109881,BMS184476, vinflunine, cryptophycin,2,3,4,5,6-pentafluoro-N-(3-fluoro-4-methoxyphenyl) benzene sulfonamide,anhydrovinblastine, TDX258, the epothilones (see for example U.S. Pat.Nos. 6,284,781 and 6,288,237) and BMS188797. In an example theepothilones are not included in the microtubuleinhibitors/microtubule-stabilising agents.

Some examples of topoisomerase inhibitors are topotecan, hycaptamine,irinotecan, rubitecan,6-ethoxypropionyl-3′,4′-O-exo-benzylidene-chartreusin,9-methoxy-N,N-dimethyl-5-nitropyrazolo[3,4,5-kl]acridine-2-(6H)propanamine,1-amino-9-ethyl-5-fluoro-2,3-dihydro-9-hydroxy-4-methyl-1H,12H-benzo[de]pyrano[3′,4′:b,7]-indolizino[1,2b]quinoline-10,13(9H,15H)dione,lurtotecan, 7-[2-(N-isopropylamino)ethyl]-(20S)camptothecin, BNP1350,BNPI1100, BN80915, BN80942, etoposide phosphate, teniposide, sobuzoxane,2′-dimethylamino-2′-deoxy-etoposide, GL331,N-[2-(dimethylamino)ethyl]-9-hydroxy-5,6-dimethyl-6H-pyrido[4,3-b]carbazole-1-carboxamide,asulacrine,(5a,5aB,8aa,9b)-9-[2-[N-[2-(dimethylamino)ethyl]-N-methylamino]ethyl]-5-[4-hydroOxy-3,5-dimethoxyphenyl]-5,5a,6,8,8a,9-hexohydrofuro(3′,4′:6,7)naphtho(2,3-d)-1,3-dioxol-6-one,2,3-(methylenedioxy)-5-methyl-7-hydroxy-8-methoxybenzo[c]-phenanthridinium,6,9-bis[(2-aminoethyl)amino]benzo[g]isoguinoline-5,10-dione,5-(3-aminopropylamino)-7,10-dihydroxy-2-(2-hydroxyethylaminomethyl)-6H-pyrazolo[4,5,1-de]acridin-6-one,N-[1-[2(diethylamino)ethylamino]-7-methoxy-9-oxo-9H-thioxanthen-4-ylmethyl]formamide,N-(2-(dimethylamino)ethyl)acridine-4-carboxamide,6-[[2-(dimethylamino)ethyl]amino]-3-hydroxy-7H-indeno[2,1-c]quinolin-7-one, and dimesna.

Examples of inhibitors of mitotic kinesins, and in particular the humanmitotic kinesin KSP, are described in Publications WO03/039460,WO03/050064, WO03/050122, WO03/049527, WO03/049679, WO03/049678,WO04/039774, WO03/079973, WO03/099211, WO03/105855, WO03/106417,WO04/037171, WO04/058148, WO04/058700, WO04/126699, WO05/018638,WO05/019206, WO05/019205, WO05/018547, WO05/017190, US2005/0176776. Inan example inhibitors of mitotic kinesins include, but are not limitedto inhibitors of KSP, inhibitors of MKLP1, inhibitors of CENP-E,inhibitors of MCAK and inhibitors of Rab6-KIFL.

Examples of “histone deacetylase inhibitors” include, but are notlimited to, SAHA, TSA, oxamflatin, PXD101, MG98 and scriptaid. Furtherreference to other histone deacetylase inhibitors may be found in thefollowing manuscript; Miller, T. A. et al. J. Med. Chem.46(24):5097-5116 (2003).

“Inhibitors of kinases involved in mitotic progression” include, but arenot limited to, inhibitors of aurora kinase, inhibitors of Polo-likekinases (PLK; in particular inhibitors of PLK-1), inhibitors of bub-1and inhibitors of bub-R1. An example of an “aurora kinase inhibitor” isVX-680 (tozasertib).

“Antiproliferative agents” includes antisense RNA and DNAoligonucleotides such as G3139, ODN698, GEM231, and INX3001, andantimetabolites such as enocitabine, carmofur, tegafur, pentostatin,doxifluridine, trimetrexate, fludarabine, capecitabine, galocitabine,cytarabine ocfosfate, fosteabine sodium hydrate, raltitrexed,paltitrexid, emitefur, tiazofurin, decitabine, nolatrexed, pemetrexed,nelzarabine, 2′-deoxy-2′-methylidenecytidine,2′-fluoromethylene-2′-deoxycytidine,N-[5-(2,3-dihydro-benzofuryl)sulfonyl]-N′-(3,4-dichlorophenyl)urea,N6-[4-deoxy-4-[N2-[2(E),4(E)-tetradecadienoyl]glycylamino]-L-glycero-B-L-manno-heptopyranosyl]adenine,aplidine, ecteinascidin, troxacitabine,4-[2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimidino[5,4-b][1,4]thiazin-6-yl-(S)-ethyl]-2,5-thienoyl-L-glutamicacid, aminopterin, 5-flurouracil, alanosine,11-acetyl-8-(carbamoyloxymethyl)-4-formyl-6-methoxy-14-oxa-1,11-diazatetracyclo(7.4.1.0.0)-tetradeca-2,4,6-trien-9-ylacetic acid ester, swainsonine, lometrexol, dexrazoxane, methioninase,2′-cyano-2′-deoxy-N4-palmitoyl-1-B-D-arabino furanosyl cytosine,3-aminopyridine-2-carboxaldehyde thiosemicarbazone and trastuzumab.

Examples of monoclonal antibody targeted therapeutic agents includethose therapeutic agents which have cytotoxic agents or radioisotopesattached to a cancer cell specific or target cell specific monoclonalantibody. Examples include Bexxar.

“HMG-CoA reductase inhibitors” refers to inhibitors of3-hydroxy-3-methylglutaryl-CoA reductase. Examples of HMG-CoA reductaseinhibitors that may be used include but are not limited to lovastatin(MEVACOR®; see U.S. Pat. Nos. 4,231,938, 4,294,926 and 4,319,039),simvastatin (ZOCOR®; see U.S. Pat. Nos. 4,444,784, 4,820,850 and4,916,239), pravastatin (PRAVACHOL®; see U.S. Pat. Nos. 4,346,227,4,537,859, 4,410,629, 5,030,447 and 5,180,589), fluvastatin (LESCOL®;see U.S. Pat. Nos. 5,354,772, 4,911,165, 4,929,437, 5,189,164,5,118,853, 5,290,946 and 5,356,896), atorvastatin (LIPITOR®; see U.S.Pat. Nos. 5,273,995, 4,681,893, 5,489,691 and 5,342,952), rosuvastatin(CRESTOR® U.S. Reissue Pat. RE37,314) and cerivastatin (also known asrivastatin and BAYCHOL®; see U.S. Pat. No. 5,177,080). The structuralformulas of these and additional HMG-CoA reductase inhibitors that maybe used in the instant methods are described at page 87 of M. Yalpani,“Cholesterol Lowering Drugs”, Chemistry & Industry, pp. 85-89 (5 Feb.1996) and U.S. Pat. Nos. 4,782,084 and 4,885,314. The term HMG-CoAreductase inhibitor as used herein includes all pharmaceuticallyacceptable lactone and open-acid forms (i.e., where the lactone ring isopened to form the free acid) as well as salt and ester forms ofcompounds which have HMG-CoA reductase inhibitory activity, and thereforthe use of such salts, esters, open-acid and lactone forms is includedwithin the scope of the invention.

“Prenyl-protein transferase inhibitor” refers to a compound whichinhibits any one or any combination of the prenyl-protein transferaseenzymes, including farnesyl-protein transferase (FPTase),geranylgeranyl-protein transferase type I (GGPTase-I), andgeranylgeranyl-protein transferase type-II (GGPTase-II, also called RabGGPTase).

Examples of prenyl-protein transferase inhibitors can be found in thefollowing publications and patents: WO 96/30343, WO 97/18813, WO97/21701, WO 97/23478, WO 97/38665, WO 98/28980, WO 98/29119, WO95/32987, U.S. Pat. Nos. 5,420,245, 5,523,430, 5,532,359, 5,510,510,5,589,485, and 5,602,098, European Patent Publ. 0 618 221, EuropeanPatent Publ. 0 675 112, European Patent Publ. 0 604 181, European PatentPubl. 0 696 593, WO 94/19357, WO 95/08542, WO 95/11917, WO 95/12612, WO95/12572, WO 95/10514, U.S. Pat. No. 5,661,152, WO 95/10515, WO95/10516, WO 95/24612, WO 95/34535, WO 95/25086, WO 96/05529, WO96/06138, WO 96/06193, WO 96/16443, WO 96/21701, WO 96/21456, WO96/22278, WO 96/24611, WO 96/24612, WO 96/05168, WO 96/05169, WO96/00736, U.S. Pat. No. 5,571,792, WO 96/17861, WO 96/33159, WO96/34850, WO 96/34851, WO 96/30017, WO 96/30018, WO 96/30362, WO96/30363, WO 96/31111, WO 96/31477, WO 96/31478, WO 96/31501, WO97/00252, WO 97/03047, WO 97/03050, WO 97/04785, WO 97/02920, WO97/17070, WO 97/23478, WO 97/26246, WO 97/30053, WO 97/44350, WO98/02436, and U.S. Pat. No. 5,532,359. For an example of the role of aprenyl-protein transferase inhibitor on angiogenesis see European J. ofCancer, Vol. 35, No. 9, pp. 1394-1401 (1999).

“Angiogenesis inhibitors” refers to compounds that inhibit the formationof new blood vessels, regardless of mechanism. Examples of angiogenesisinhibitors include, but are not limited to, tyrosine kinase inhibitors,such as inhibitors of the tyrosine kinase receptors Flt-1 (VEGFR1) andFlk-1/KDR (VEGFR2), inhibitors of epidermal-derived, fibroblast-derived,or platelet derived growth factors, MMP (matrix metalloprotease)inhibitors, integrin blockers, interferon-a, interleukin-12, pentosanpolysulfate, cyclooxygenase inhibitors, including nonsteroidalanti-inflammatories (NSAIDs) like aspirin and ibuprofen as well asselective cyclooxy-genase-2 inhibitors like celecoxib and rofecoxib(PNAS, Vol. 89, p. 7384 (1992); JNCI, Vol. 69, p. 475 (1982); Arch.Opthalmol., Vol. 108, p. 573 (1990); Anat. Rec., Vol. 238, p. 68 (1994);FEBS Letters, Vol. 372, p. 83 (1995); Clin, Orthop. Vol. 313, p. 76(1995); J. Mol. Endocrinol., Vol. 16, p. 107 (1996); Jpn. J. Pharmacol.,Vol. 75, p. 105 (1997); Cancer Res., Vol. 57, p. 1625 (1997); Cell, Vol.93, p. 705 (1998); Intl. J. Mol. Med., Vol. 2, p. 715 (1998); J. Biol.Chem., Vol. 274, p. 9116 (1999)), steroidal anti-inflammatories (such ascorticosteroids, mineralocorticoids, dexamethasone, prednisone,prednisolone, methylpred, betamethasone), carboxyamidotriazole,combretastatin A-4, squalamine, 6-O-chloroacetyl-carbonyl)-fumagillol,thalidomide, angiostatin, troponin-1, angiotensin II antagonists (seeFernandez et al., J. Lab. Clin. Med. 105:141-145 (1985)), and antibodiesto VEGF (see, Nature Biotechnology, Vol. 17, pp. 963-968 (October 1999);Kim et al., Nature, 362, 841-844 (1993); WO 00/44777; and WO 00/61186).

Other therapeutic agents that modulate or inhibit angiogenesis and mayalso be used in combination with the compounds of the instant inventioninclude agents that modulate or inhibit the coagulation and fibrinolysissystems (see review in Clin. Chem. La. Med. 38:679-692 (2000)). Examplesof such agents that modulate or inhibit the coagulation and fibrinolysispathways include, but are not limited to, heparin (see Thromb. Haemost.80:10-23 (1998)), low molecular weight heparins and carboxypeptidase Uinhibitors (also known as inhibitors of active thrombin activatablefibrinolysis inhibitor [TAFIa]) (see Thrombosis Res. 101:329-354(2001)). TAFIa inhibitors have been described in U.S. Ser. Nos.60/310,927 (filed Aug. 8, 2001) and 60/349,925 (filed Jan. 18, 2002).

“Agents that interfere with cell cycle checkpoints” refer to compoundsthat inhibit protein kinases that transduce cell cycle checkpointsignals, thereby sensitizing the cancer cell to DNA damaging agents.Such agents include inhibitors of ATR, ATM, the CHK1 and CHK2 kinasesand cdk and cdc kinase inhibitors and are specifically exemplified by7-hydroxystaurosporin, flavopiridol, CYC202 (Cyclacel) and BMS-387032.

“Agents that interfere with receptor tyrosine kinases (RTKs)” refer tocompounds that inhibit RTKs and therefore mechanisms involved inoncogenesis and tumor progression. Such agents include inhibitors ofc-Kit, Eph, PDGF, Flt3 and c-Met. Further agents include inhibitors ofRTKs as described by Bume-Jensen and Hunter, Nature, 411:355-365, 2001.

“Inhibitors of cell proliferation and survival signalling pathway” referto compounds that inhibit signal transduction cascades downstream ofcell surface receptors. Such agents include inhibitors ofserine/threonine kinases (including but not limited to inhibitors of Aktsuch as described in WO 02/083064, WO 02/083139, WO 02/083140, US2004-0116432, WO 02/083138, US 2004-0102360, WO 03/086404, WO 03/086279,WO 03/086394, WO 03/084473, WO 03/086403, WO 2004/041162, WO2004/096131, WO 2004/096129, WO 2004/096135, WO 2004/096130, WO2005/100356, WO 2005/100344, US 2005/029941, US 2005/44294, US2005/43361, 60/734188, 60/652737, 60/670469), inhibitors of Raf kinase(for example BAY-43-9006), inhibitors of MEK (for example CI-1040 andPD-098059), inhibitors of mTOR (for example Wyeth CCI-779), andinhibitors of PI3K (for example LY294002).

As described above, the combinations with NSAID's are directed to theuse of NSAID's which are potent COX-2 inhibiting agents. For purposes ofthe specification an NSAID is potent if it possesses an IC₅₀ for theinhibition of COX-2 of 1 μM or less as measured by cell or microsomalassays.

The invention also encompasses combinations with NSAID's which areselective COX-2 inhibitors. For purposes of the specification NSAID'swhich are selective inhibitors of COX-2 are defined as those whichpossess a specificity for inhibiting COX-2 over COX-1 of at least 100fold as measured by the ratio of IC₅₀ for COX-2 over IC₅₀ for COX-1evaluated by cell or microsomal assays. Such compounds include, but arenot limited to those disclosed in U.S. Pat. Nos. 5,474,995, 5,861,419,6,001,843, 6,020,343, 5,409,944, 5,436,265, 5,536,752, 5,550,142,5,604,260, 5,698,584, 5,710,140, WO 94/15932, U.S. Pat. Nos. 5,344,991,5,134,142, 5,380,738, 5,393,790, 5,466,823, 5,633,272 and U.S. Pat. No.5,932,598, all of which are hereby incorporated by reference.

Inhibitors of COX-2 that are particularly useful in the instant methodof treatment are: 3-phenyl-4-(4-(methylsulfonyl)phenyl)-2-(5H)-furanone;and5-chloro-3-(4-methylsulfonyl)-phenyl-2-(2-methyl-5-pyridinyl)pyridine;or a pharmaceutically acceptable salt thereof.

Compounds that have been described as specific inhibitors of COX-2 andare therefore useful in the present invention include, but are notlimited to, the following: parecoxib, BEXTRA® and CELEBREX® or apharmaceutically acceptable salt thereof.

Other examples of angiogenesis inhibitors include, but are not limitedto, endostatin, ukrain, ranpirnase, IM862,5-methoxy-4-[2-methyl-3-(3-methyl-2-butenyl)oxiranyl]-1-oxaspiro[2,5]oct-6-yl(chloroacetyl)carbamate,acetyldinanaline,5-amino-1-[[3,5-dichloro-4-(4-chlorobenzoyl)phenyl]methyl]-1H-1,2,3-triazole-4-carboxamide,CM101,squalamine, combretastatin, RPI4610, NX31838, sulfated mannopentaosephosphate,7,7-(carbonyl-bis[imino-N-methyl-4,2-pyrrolocarbonylimino[N-methyl-4,2-pyrrole]-carbonylimino]-bis-(1,3-naphthalenedisulfonate), and 3-[(2,4-dimethylpyrrol-5-yl)methylene]-2-indolinone(SU5416).

As used above, “integrin blockers” refers to compounds which selectivelyantagonize, inhibit or counteract binding of a physiological ligand tothe α_(v)β₃ integrin, to compounds which selectively antagonize, inhibitor counteract binding of a physiological ligand to the α_(v)β₅ integrin,to compounds which antagonize, inhibit or counteract binding of aphysiological ligand to both the α_(v)β₃ integrin and the α_(v)β₅integrin, and to compounds which antagonize, inhibit or counteract theactivity of the particular integrin(s) expressed on capillaryendothelial cells. The term also refers to antagonists of the α_(v)β₆,α_(v)β₈, α₁β₁, α₂β₁, α₅β₁, α_(v)β₁ and α₆β₄ integrins. The term alsorefers to antagonists of any combination of α_(v)β₃, α_(v)β₅, α_(v)β₆,α_(v)β₈, α₁β₁, α₂β₁, α₅β₁, α₁β₁ and α₆β₄ integrins.

Some specific examples of tyrosine kinase inhibitors includeN-(trifluoromethylphenyl)-5-methylisoxazol-4-carboxamide,3-[(2,4-dimethylpyrrol-5-yl)methylidenyl)indolin-2-one,17-(allylamino)-17-demethoxygeldanamycin,4-(3-chloro-4-fluorophenylamino)-7-methoxy-6-[3-(4-morpholinyl)propoxyl]quinazoline,N-(3-ethynylphenyl)-6,7-bis(2-methoxyethoxy)-4-quinazolinamine,BIBX1382,2,3,9,10,11,12-hexahydro-10-(hydroxymethyl)-10-hydroxy-9-methyl-9,12-epoxy-1H-diindolo[1,2,3-fg:3′,2′,1′-kl]pyrrolo[3,4-i][1,6]benzodiazocin-1-one,SH268, genistein, STI571, CEP2563,4-(3-chlorophenylamino)-5,6-dimethyl-7H-pyrrolo[2,3-d]pyrimidinemethanesulfonate, 4-(3-bromo-4-hydroxyphenyl)amino-6,7-dimethoxyquinazoline,4-(4′-hydroxyphenyl)amino-6,7-dimethoxyquinazoline, SU6668, STI571A,N-4-chlorophenyl-4-(4-pyridylmethyl)-1-phthalazinamine, and EMD121974.

Combinations with compounds other than anti-cancer compounds are alsoencompassed in the instant methods. For example, combinations of theinstantly claimed compounds with PPAR-γ (i.e., PPAR-gamma) agonists andPPAR-δ (i.e., PPAR-delta) agonists are useful in the treatment ofcertain malingnancies. PPAR-γ and PPAR-δ are the nuclear peroxisomeproliferator-activated receptors γ and δ. The expression of PPAR-γ onendothelial cells and its involvement in angiogenesis has been reportedin the literature (see J. Cardiovasc. Pharmacol. 1998; 31:909-913; J.Biol. Chem. 1999; 274:9116-9121; Invest. Ophthalmol Vis. Sci. 2000;41:2309-2317). More recently, PPAR-γ agonists have been shown to inhibitthe angiogenic response to VEGF in vitro; both troglitazone androsiglitazone maleate inhibit the development of retinalneovascularization in mice. (Arch. Ophthamol. 2001; 119:709-717).Examples of PPAR-γ agonists and PPAR-γ/α agonists include, but are notlimited to, thiazolidinediones (such as DRF2725, CS-011, troglitazone,rosiglitazone, and pioglitazone), fenofibrate, gemfibrozil, clofibrate,GW2570, SB219994, AR-H039242, JTT-501, MCC-555, GW2331, GW409544,NN2344, KRP297, NP0110, DRF4158, NN622, G262570, PNU182716, DRF552926,2-[(5,7-dipropyl-3-trifluoromethyl-1,2-benzisoxazol-6-yl)oxy]-2-methylpropionicacid (disclosed in U.S. Ser. No. 09/782,856), and2(R)-7-(3-(2-chloro-4-(4-fluorophenoxy)phenoxy)propoxy)-2-ethylchromane-2-carboxylic acid (disclosed in U.S.Ser. No. 60/235,708 and 60/244,697).

Another example of the instant invention is the use of the presentlydisclosed compounds in combination with gene therapy for the treatmentof cancer. For an overview of genetic strategies to treating cancer seeHall et al (Am. J. Hum. Genet. 61:785-789, 1997) and Kufe et al (CancerMedicine, 5th Ed, pp 876-889, BC Decker, Hamilton 2000). Gene therapycan be used to deliver any tumor suppressing gene. Examples of suchgenes include, but are not limited to, p53, which can be delivered viarecombinant virus-mediated gene transfer (see U.S. Pat. No. 6,069,134,for example), a uPA/uPAR antagonist (“Adenovirus-Mediated Delivery of auPA/uPAR Antagonist Suppresses Angiogenesis-Dependent Tumor Growth andDissemination in Mice,” Gene Therapy, August 1998; 5(8):1105-13), andinterferon gamma (J. Immunol. 2000; 164:217-222).

The compounds of the instant invention may also be administered incombination with an inhibitor of inherent multidrug resistance (MDR), inparticular MDR associated with high levels of expression of transporterproteins. Such MDR inhibitors include inhibitors of p-glycoprotein(P-gp), such as LY335979, XR9576, OC144-093, R101922, VX853 and PSC833(valspodar).

A compound of the present invention may be employed in conjunction withanti-emetic agents to treat nausea or emesis, including acute, delayed,late-phase, and anticipatory emesis, which may result from the use of acompound of the present invention, alone or with radiation therapy. Forthe prevention or treatment of emesis, a compound of the presentinvention may be used in conjunction with other anti-emetic agents,especially neurokinin-1 receptor antagonists, 5HT3 receptor antagonists,such as ondansetron, granisetron, tropisetron, and zatisetron, GABABreceptor agonists, such as baclofen, a corticosteroid such as Decadron(dexamethasone), Kenalog, Aristocort, Nasalide, Preferid, Benecorten orothers such as disclosed in U.S. Pat. Nos. 2,789,118, 2,990,401,3,048,581, 3,126,375, 3,929,768, 3,996,359, 3,928,326 and 3,749,712, anantidopaminergic, such as the phenothiazines (for exampleprochlorperazine, fluphenazine, thioridazine and mesoridazine),metoclopramide or dronabinol. In another example, conjunctive therapywith an anti-emesis agent selected from a neurokinin-1 receptorantagonist, a 5HT3 receptor antagonist and a corticosteroid is disclosedfor the treatment or prevention of emesis that may result uponadministration of the instant compounds.

Neurokinin-1 receptor antagonists of use in conjunction with thecompounds of the present invention are fully described, for example, inU.S. Pat. Nos. 5,162,339, 5,232,929, 5,242,930, 5,373,003, 5,387,595,5,459,270, 5,494,926, 5,496,833, 5,637,699, 5,719,147; European PatentPublication Nos. EP 0 360 390, 0 394 989, 0 428 434, 0 429 366, 0 430771, 0 436334,0443 132,0482539,0498069,0499313,0512901,0512902,0514273,0514 274,0514275,0514276,0515681,0517589,0520555,0522808,0528495,0532456,0 533 280, 0 536 817, 0 545478, 0 558 156, 0 577 394, 0 585 913,0 590 152, 0 599 538, 0 610793,0634402,0686629,0693489,0694535,0699655,0699674,0707006,0708 101,0709 375, 0 709 376, 0 714 891, 0 723 959, 0 733 632 and 0 776 893; PCTInternational Patent Publication Nos. WO 90/05525, 90/05729, 91/09844,91/18899, 92/01688, 92/06079, 92/12151, 92/15585, 92/17449, 92/20661,92/20676, 92/21677, 92/22569, 93/00330, 93/00331, 93/01159,93/01165,93/01169,93/01170,93/06099,93/09116,93/10073,93/14084,93/14113,93/18023,93/19064, 93/21155, 93/21181, 93/23380, 93/24465, 94/00440, 94/01402,94/02461, 94/02595, 94/03429, 94/03445, 94/04494, 94/04496, 94/05625,94/07843, 94/08997, 94/10165, 94/10167, 94/10168,94/10170,94/11368,94/13639, 94/13663,94/14767, 94/15903,94/19320, 94/19323, 94/20500,94/26735, 94/26740, 94/29309, 95/02595, 95/04040, 95/04042, 95/06645,95/07886, 95/07908,95/08549,95/11880, 95/14017, 95/15311, 95/16679,95/17382,95/18124, 95/18129, 95/19344, 95/20575, 95/21819, 95/22525,95/23798, 95/26338, 95/28418, 95/30674, 95/30687, 95/33744, 96/05181,96/05193, 96/05203, 96/06094, 96/07649, 96/10562, 96/16939, 96/18643,96/20197, 96/21661, 96/29304, 96/29317, 96/29326, 96/29328, 96/31214,96/32385, 96/37489, 97/01553, 97/01554, 97/03066, 97/08144, 97/14671,97/17362, 97/18206, 97/19084, 97/19942 and 97/21702; and in BritishPatent Publication Nos. 2 266 529, 2 268 931, 2 269 170, 2 269 590, 2271774, 2 292 144, 2 293 168, 2 293 169, and 2 302 689. The preparationof such compounds is fully described in the aforementioned patents andpublications, which are incorporated herein by reference.

In an example, the neurokinin-1 receptor antagonist for use inconjunction with the compounds of the present invention is selectedfrom:2-(R)-(1-(R)-(3,5-bis(trifluoromethyl)phenyl)ethoxy)-3-(S)-(4-fluorophenyl)-4-(3-(5-oxo-1H,4H-1,2,4-triazolo)methyl)morpholine,or a pharmaceutically acceptable salt thereof, which is described inU.S. Pat. No. 5,719,147.

A compound of the instant invention may also be administered with anagent useful in the treatment of anemia. Such an anemia treatment agentis, for example, a continuous erythropoiesis receptor activator (such asepoetin alfa).

A compound of the instant invention may also be administered with anagent useful in the treatment of neutropenia. Such a neutropeniatreatment agent is, for example, a hematopoietic growth factor whichregulates the production and function of neutrophils such as a humangranulocyte colony stimulating factor, (G-CSF). Examples of a G-CSFinclude filgrastim.

A compound of the instant invention may also be administered with animmunologic-enhancing drug, such as levamisole, isoprinosine andZadaxin.

A compound of the instant invention may also be useful for treating orpreventing cancer in combination with P450 inhibitors including:xenobiotics, quinidine, tyramine, ketoconazole, testosterone, quinine,methyrapone, caffeine, phenelzine, doxorubicin, troleandomycin,cyclobenzaprine, erythromycin, cocaine, furafyline, cimetidine,dextromethorphan, ritonavir, indinavir, amprenavir, diltiazem,terfenadine, verapamil, cortisol, itraconazole, mibefradil, nefazodoneand nelfinavir.

A compound of the instant invention may also be useful for treating orpreventing cancer in combination with Pgp and/or BCRP inhibitorsincluding: cyclosporin A, PSC833, GF120918, cremophorEL, fumitremorginC, Ko132, Ko134, Iressa, Imatnib mesylate, EKI-785, C11033, novobiocin,diethylstilbestrol, tamoxifen, resperpine, VX-710, tryprostatin A,flavonoids, ritonavir, saquinavir, nelfinavir, omeprazole, quinidine,verapamil, terfenadine, ketoconazole, nifidepine, FK506, amiodarone,XR9576, indinavir, amprenavir, cortisol, testosterone, LY335979,OC144-093, erythromycin, vincristine, digoxin and talinolol.

A compound of the instant invention may also be useful for treating orpreventing cancer, including bone cancer, in combination withbisphosphonates (understood to include bisphosphonates, diphosphonates,bisphosphonic acids and diphosphonic acids). Examples of bisphosphonatesinclude but are not limited to: etidronate (Didronel), pamidronate(Aredia), alendronate (Fosamax), risedronate (Actonel), zoledronate(Zometa), ibandronate (Boniva), incadronate or cimadronate, clodronate,EB-1053, minodronate, neridronate, piridronate and tiludronate includingany and all pharmaceutically acceptable salts, derivatives, hydrates andmixtures thereof.

A compound of the instant invention may also be useful for treating orpreventing breast cancer in combination with aromatase inhibitors.Examples of aromatase inhibitors include but are not limited to:anastrozole, letrozole and exemestane.

A compound of the instant invention may also be useful for treating orpreventing cancer in combination with siRNA therapeutics.

The compounds of the instant invention may also be administered incombination with γ-secretase inhibitors and/or inhibitors of NOTCHsignaling. Such inhibitors include compounds described in WO 01/90084,WO 02/30912, WO 01/70677, WO 03/013506, WO 02/36555, WO 03/093252, WO03/093264, WO 03/093251, WO 03/093253, WO 2004/039800, WO 2004/039370,WO 2005/030731, WO 2005/014553, U.S. Ser. No. 10/957,251, WO2004/089911, WO 02/081435, WO 02/081433, WO 03/018543, WO 2004/031137,WO 2004/031139, WO 2004/031138, WO 2004/101538, WO 2004/101539 and WO02/47671 (including LY-450139).

A compound of the instant invention may also be useful for treating orpreventing cancer in combination with PARP inhibitors.

A compound of the instant invention may also be useful for treatingcancer in combination with the following therapeutic agents:pembrolizumab (Keytruda®), abarelix (Plenaxis Depot®); aldesleukin(Prokine®); Aldesleukin (Proleukin®); Alemtuzumabb (Campath®);alitretinoin (Panretin®); allopurinol (Zyloprim®); altretamine(Hexalen®); amifostine (Ethyol®); anastrozole (Arimidex®); arsenictrioxide (Trisenox®); asparaginase (Elspar®); azacitidine (Vidaza®);bevacuzimab (Avastin®); bexarotene capsules (Targretin®); bexarotene gel(Targretin®); bleomycin (Blenoxane®); bortezomib (Velcade®); busulfanintravenous (Busulfex®); busulfan oral (Myleran®); calusterone(Methosarb®); capecitabine (Xeloda®); carboplatin (Paraplatin®);carmustine (BCNU®, BiCNU®); carmustine (Gliadel®); carmustine withPolifeprosan 20 Implant (Gliadel Wafer®); celecoxib (Celebrex®);cetuximab (Erbitux®); chlorambucil (Leukeran®); cisplatin (Platinol®);cladribine (Leustatin®, 2-CdA®); clofarabine (Clolar®); cyclophosphamide(Cytoxan®, Neosar®); cyclophosphamide (Cytoxan Injection®);cyclophosphamide (Cytoxan Tablet®); cytarabine (Cytosar-U®); cytarabineliposomal (DepoCyt®); dacarbazine (DTIC-Dome®); dactinomycin,actinomycin D (Cosmegen®); Darbepoetin alfa (Aranesp®); daunorubicinliposomal (DanuoXome®); daunorubicin, daunomycin (Daunorubicin®);daunorubicin, daunomycin (Cerubidine®); Denileukin diftitox (Ontak®);dexrazoxane (Zinecard®); docetaxel (Taxotere®); doxorubicin (AdriamycinPFS®); doxorubicin (Adriamycin®, Rubex®); doxorubicin (Adriamycin PFSInjection®); doxorubicin liposomal (Doxil®); dromostanolone propionate(Dromostanolone®); dromostanolone propionate (Masterone Injection®);Elliott's B Solution (Elliott's B Solution®); epirubicin (Ellence®);Epoetin alfa (Epogen®); erlotinib (Tarceva®); estramustine (Emcyt®);etoposide phosphate (Etopophos®); etoposide, VP-16 (Vepesid®);exemestane (Aromasin®); Filgrastim (Neupogen®); floxuridine(intraarterial) (FUDR®); fludarabine (Fludara®); fluorouracil, 5-FU(Adrucil®); fulvestrant (Faslodex®); gefitinib (Iressa®); gemcitabine(Gemzar®); gemtuzumab ozogamicin (Mylotarg®); goserelin acetate (ZoladexImplant®); goserelin acetate (Zoladex®); histrelin acetate (HistrelinImplant®); hydroxyurea (Hydrea®); Ibritumomab Tiuxetan (Zevalin®);idarubicin (Idamycin®); ifosfamide (IFEX®); imatinib mesylate(Gleevec®); interferon alfa 2a (Roferon A®); Interferon alfa-2b (IntronA®); irinotecan (Camptosar®); lenalidomide (Reylimid®); letrozole(Femara®); leucovorin (Wellcovorin®, Leucovorin®); Leuprolide Acetate(Eligard®); levamisole (Ergamisol®); lomustine, CCNU (CeeBU®);meclorethamine, nitrogen mustard (Mustargen®); megestrol acetate(Megace®); melphalan, L-PAM (Alkeran®); mercaptopurine, 6-MP(Purinethol®); mesna (Mesnex®); mesna (Mesnex Tabs®); methotrexate(Methotrexate®); methoxsalen (Uvadex®); mitomycin C (Mutamycin®);mitotane (Lysodren®); mitoxantrone (Novantrone®); nandrolonephenpropionate (Durabolin-50@); nelarabine (Arranon®); Nofetumomab(Verluma®); Oprelvekin (Neumega®); oxaliplatin (Eloxatin®); paclitaxel(Paxene®); paclitaxel (Taxol®); paclitaxel protein-bound particles(Abraxane®); palifermin (Kepivance®); pamidronate (Aredia®); pegademase(Adagen (Pegademase Bovine)®); pegaspargase (Oncaspar®); Pegfilgrastim(Neulasta®); pemetrexed disodium (Alimta®); pentostatin (Nipent®);pipobroman (Vercyte®); plicamycin, mithramycin (Mithracin®); porfimersodium (Photofrin®); procarbazine (Matulane®); quinacrine (Atabrine®);Rasburicase (Elitek®); Rituximab (Rituxan®); Ridaforolimus; sargramostim(Leukine®); Sargramostim (Prokine®); sorafenib (Nexavar®); streptozocin(Zanosar®); sunitinib maleate (Sutent®); talc (Sclerosol®); tamoxifen(Nolvadex®); temozolomide (Temodar®); teniposide, VM-26 (Vumon®);testolactone (Teslac®); thioguanine, 6-TG (Thioguanine®); thiotepa(Thioplex®); topotecan (Hycamtin®); toremifene (Fareston®); Tositumomab(Bexxar®); Tositumomab/I-131 tositumomab (Bexxar®); Trastuzumab(Herceptin®); tretinoin, ATRA (Vesanoid®); Uracil Mustard (UracilMustard Capsules®); valrubicin (Valstar®); vinblastine (Velban®);vincristine (Oncovin®); vinorelbine (Navelbine®); vorinostat (Zolinza®)and zoledronate (Zometa®).

In an example, the angiogenesis inhibitor to be used as the secondcompound is selected from a tyrosine kinase inhibitor, an inhibitor ofepidermal-derived growth factor, an inhibitor of fibroblast-derivedgrowth factor, an inhibitor of platelet derived growth factor, an MMP(matrix metalloprotease) inhibitor, an integrin blocker, interferon-a,interleukin-12, pentosan polysulfate, a cyclooxygenase inhibitor,carboxyamidotriazole, combretastatin A-4, squalamine,6-O-chloroacetyl-carbonyl)-fumagillol, thalidomide, angiostatin,troponin-1, or an antibody to VEGF. In an example, the estrogen receptormodulator is tamoxifen or raloxifene.

Thus, the scope of the instant invention encompasses the use of theinstantly claimed compounds in combination with a second compoundselected from: an estrogen receptor modulator, an androgen receptormodulator, a retinoid receptor modulator, a cytotoxic/cytostatic agent,an antiproliferative agent, a prenyl-protein transferase inhibitor, anHMG-CoA reductase inhibitor, an HIV protease inhibitor, a reversetranscriptase inhibitor, an angiogenesis inhibitor, PPAR-γ agonists,PPAR-δ agonists, an inhibitor of inherent multidrug resistance, ananti-emetic agent, an agent useful in the treatment of anemia, an agentuseful in the treatment of neutropenia, an immunologic-enhancing drug,an inhibitor of cell proliferation and survival signaling, abisphosphonate, an aromatase inhibitor, an siRNA therapeutic,γ-secretase and/or NOTCH inhibitors, agents that interfere with receptortyrosine kinases (RTKs), an agent that interferes with a cell cyclecheckpoint, and any of the therapeutic agents listed above.

Also included in the scope of the claims is a method of treating cancerthat comprises administering a therapeutically effective amount of acompound of the instant invention in combination with radiation therapyand/or in combination with a second compound selected from: an estrogenreceptor modulator, an androgen receptor modulator, a retinoid receptormodulator, a cytotoxiccytostatic agent, an antiproliferative agent, aprenyl-protein transferase inhibitor, an HMG-CoA reductase inhibitor, anHIV protease inhibitor, a reverse transcriptase inhibitor, anangiogenesis inhibitor, PPAR-γ agonists, PPAR-δ agonists, an inhibitorof inherent multidrug resistance, an anti-emetic agent, an agent usefulin the treatment of anemia, an agent useful in the treatment ofneutropenia, an immunologic-enhancing drug, an inhibitor of cellproliferation and survival signaling, a bisphosphonate, an aromataseinhibitor, an siRNA therapeutic, γ-secretase and/or NOTCH inhibitors,agents that interfere with receptor tyrosine kinases (RTKs), an agentthat interferes with a cell cycle checkpoint, and any of the therapeuticagents listed above.

And yet another example of the invention is a method of treating cancerthat comprises administering a therapeutically effective amount of acompound of the instant invention in combination with paclitaxel ortrastuzumab.

The invention further encompasses a method of treating or preventingcancer that comprises administering a therapeutically effective amountof a compound of the instant invention in combination with a COX-2inhibitor.

The instant invention also includes a pharmaceutical composition usefulfor treating or preventing cancer that comprises a therapeuticallyeffective amount of a compound of the instant invention and a secondcompound selected from: an estrogen receptor modulator, an androgenreceptor modulator, a retinoid receptor modulator, acytotoxic/cytostatic agent, an antiproliferative agent, a prenyl-proteintransferase inhibitor, an HMG-CoA reductase inhibitor, an HIV proteaseinhibitor, a reverse transcriptase inhibitor, an angiogenesis inhibitor,a PPAR-γ agonist, a PPAR-δ agonist, an inhibitor of cell proliferationand survival signaling, a bisphosphonate, an aromatase inhibitor, ansiRNA therapeutic, γ-secretase and/or NOTCHinhibitors, agents thatinterfere with receptor tyrosine kinases (RTKs), an agent thatinterferes with a cell cycle checkpoint, and any of the therapeuticagents listed above.

When any variable occurs more than one time in any constituent, itsdefinition on each occurrence is independent at every other occurrence.Also, combinations of substituents and variables are permissible only ifsuch combinations result in stable compounds. Lines drawn into the ringsystems from substituents indicate that the indicated bond may beattached to any of the substitutable ring atoms. If the ring system isbicyclic, it is intended that the bond be attached to any of thesuitable atoms on either ring of the bicyclic moiety.

It is understood that substituents and substitution patterns on thecompounds of the instant invention can be selected by one of ordinaryskill in the art to provide compounds that are chemically stable andthat can be readily synthesized by techniques known in the art, as wellas those methods set forth below, from readily available startingmaterials. If a substituent is itself substituted with more than onegroup, it is understood that these multiple groups may be on the samecarbon or on different carbons, so long as a stable structure results.Also, “optionally substituted” means optional substitution with thespecified groups, radicals or moieties.

It will be understood that, as used herein, references to the compoundsof structural Formula I are meant to also include the pharmaceuticallyacceptable salts, and also salts that are not pharmaceuticallyacceptable when they are used as precursors to the free compounds ortheir pharmaceutically acceptable salts or in other syntheticmanipulations.

The compounds of the present invention may be administered in the formof a pharmaceutically acceptable salt. The term “pharmaceuticallyacceptable salt” refers to salts prepared from pharmaceuticallyacceptable non-toxic bases or acids including inorganic or organic basesand inorganic or organic acids. Salts of basic compounds encompassedwithin the term “pharmaceutically acceptable salt” refer to non-toxicsalts of the compounds of the invention which are generally prepared byreacting the free base with a suitable organic or inorganic acid.Representative salts of basic compounds of the present inventioninclude, but are not limited to, the following: acetate, ascorbate,adipate, alginate, aspirate, benzenesulfonate, benzoate, bicarbonate,bisulfate, bitartrate, borate, bromide, butyrate, camphorate,camphorsulfonate, camsylate, carbonate, chloride, clavulanate, citrate,cyclopentane propionate, diethylacetic, digluconate, dihydrochloride,dodecylsulfanate, edetate, edisylate, estolate, esylate,ethanesulfonate, formic, fumarate, gluceptate, glucoheptanoate,gluconate, glutamate, glycerophosphate, glycollylarsanilate,hemisulfate, heptanoate, hexanoate, hexylresorcinate, hydrabamine,hydrobromide, hydrochloride, 2-hydroxyethanesulfonate,hydroxynaphthoate, iodide, isonicotinic, isothionate, lactate,lactobionate, laurate, malate, maleate, mandelate, mesylate,methylbromide, methylnitrate, methylsulfate, methanesulfonate, mucate,2-naphthalenesulfonate, napsylate, nicotinate, nitrate,N-methylglucamine ammonium salt, oleate, oxalate, pamoate (embonate),palmitate, pantothenate, pectinate, persulfate, phosphate/diphosphate,pimelic, phenylpropionic, polygalacturonate, propionate, salicylate,stearate, sulfate, subacetate, succinate, tannate, tartrate, teoclate,thiocyanate, tosylate, triethiodide, trifluoroacetate, undeconate,valerate and the like. Furthermore, where the compounds of the inventioncarry an acidic moiety, suitable pharmaceutically acceptable saltsthereof include, but are not limited to, salts derived from inorganicbases including aluminum, ammonium, calcium, copper, ferric, ferrous,lithium, magnesium, manganic, mangamous, potassium, sodium, zinc, andthe like. Particularly preferred are the ammonium, calcium, magnesium,potassium, and sodium salts. Salts derived from pharmaceuticallyacceptable organic non-toxic bases include salts of primary, secondary,and tertiary amines, cyclic amines, dicyclohexyl amines and basicion-exchange resins, such as arginine, betaine, caffeine, choline,N,N-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol,2-dimethylaminoethanol, ethanolamine, ethylamine, ethylenediamine,N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine,hydrabamine, isopropylamine, lysine, methylglucamine, morpholine,piperazine, piperidine, polyamine resins, procaine, purines,theobromine, triethylamine, trimethylamine, tripropylamine,tromethamine, and the like. Also, included are the basicnitrogen-containing groups may be quaternized with such agents as loweralkyl halides, such as methyl, ethyl, propyl, and butyl chloride,bromides and iodides; dialkyl sulfates like dimethyl, diethyl, dibutyl;and diamyl sulfates, long chain halides such as decyl, lauryl, myristyland stearyl chlorides, bromides and iodides, aralkyl halides like benzyland phenethyl bromides and others.

These salts can be obtained by known methods, for example, by mixing acompound of the present invention with an equivalent amount and asolution containing a desired acid, base, or the like, and thencollecting the desired salt by filtering the salt or distilling off thesolvent. The compounds of the present invention and salts thereof mayform solvates with a solvent such as water, ethanol, or glycerol. Thecompounds of the present invention may form an acid addition salt and asalt with a base at the same time according to the type of substituentof the side chain.

The present invention encompasses all stereoisomeric forms of thecompounds of Formula I. Centers of asymmetry that are present in thecompounds of Formula I can all independently of one another have (R)configuration or (S) configuration. When bonds to the chiral carbon aredepicted as straight lines in the structural Formulas of the invention,it is understood that both the (R) and (S) configurations of the chiralcarbon, and hence both enantiomers and mixtures thereof, are embracedwithin the Formula. Similarly, when a compound name is recited without achiral designation for a chiral carbon, it is understood that both the(R) and (S) configurations of the chiral carbon, and hence individualenantiomers and mixtures thereof, are embraced by the name. Theproduction of specific stereoisomers or mixtures thereof may beidentified in the Examples where such stereoisomers or mixtures wereobtained, but this in no way limits the inclusion of all stereoisomersand mixtures thereof from being within the scope of the invention.

The invention includes all possible enantiomers and diastereomers andmixtures of two or more stereoisomers, for example mixtures ofenantiomers and/or diastereomers, in all ratios. Thus, enantiomers are asubject of the invention in enantiomerically pure form, both aslevorotatory and as dextrorotatory antipodes, in the form of racematesand in the form of mixtures of the two enantiomers in all ratios. In thecase of a cis/trans isomerism the invention includes both the cis formand the trans form as well as mixtures of these forms in all ratios. Thepreparation of individual stereoisomers can be carried out, if desired,by separation of a mixture by customary methods, for example bychromatography or crystallization, by the use of stereochemicallyuniform starting materials for the synthesis or by stereoselectivesynthesis. Optionally a derivatization can be carried out before aseparation of stereoisomers. The separation of a mixture ofstereoisomers can be carried out at an intermediate step during thesynthesis of a compound of Formula I or it can be done on a finalracemic product. Absolute stereochemistry may be determined by X-raycrystallography of crystalline products or crystalline intermediateswhich are derivatized, if necessary, with a reagent containing astereogenic center of known configuration. Where compounds of theinvention are capable of tautomerization, all individual tautomers aswell as mixtures thereof are included in the scope of the invention. Thepresent invention includes all such isomers, as well as salts, solvates(including hydrates) and solvated salts of such racemates, enantiomers,diastereomers and tautomers and mixtures thereof.

Centers of asymmetry that are present in the compounds of Formula I canall independently of one another have (R) configuration or (S)configuration. When bonds to the chiral carbon are depicted as straightlines in the structural Formulas of the invention, it is understood thatboth the (R) and (S) configurations of the chiral carbon, and hence bothenantiomers and mixtures thereof, are embraced within the Formula.Similarly, when a compound name is recited without a chiral designationfor a chiral carbon, it is understood that both the (R) and (S)configurations of the chiral carbon, and hence individual enantiomersand mixtures thereof, are embraced by the name. The production ofspecific stereoisomers or mixtures thereof may be identified in theExamples where such stereoisomers or mixtures were obtained, but this inno way limits the inclusion of all stereoisomers and mixtures thereoffrom being within the scope of the invention.

In the compounds of the invention, the atoms may exhibit their naturalisotopic abundances, or one or more of the atoms may be artificiallyenriched in a particular isotope having the same atomic number, but anatomic mass or mass number different from the atomic mass or mass numberpredominantly found in nature. The present invention is meant to includeall suitable isotopic variations of the specifically and genericallydescribed compounds. For example, different isotopic forms of hydrogen(H) include protium (¹H) and deuterium (²H). Protium is the predominanthydrogen isotope found in nature. Enriching for deuterium may affordcertain therapeutic advantages, such as increasing in vivo half-life orreducing dosage requirements, or may provide a compound useful as astandard for characterization of biological samples.Isotopically-enriched compounds can be prepared without undueexperimentation by conventional techniques well known to those skilledin the art or by processes analogous to those described in the generalprocess schemes and examples herein using appropriateisotopically-enriched reagents and/or intermediates.

Furthermore, compounds of the present invention may exist in amorphousform and/or one or more crystalline forms, and as such all amorphous andcrystalline forms and mixtures thereof of the compounds of Formula I areintended to be included within the scope of the present invention. Inaddition, some of the compounds of the instant invention may formsolvates with water (i.e., a hydrate) or common organic solvents. Suchsolvates and hydrates, particularly the pharmaceutically acceptablesolvates and hydrates, of the instant compounds are likewise encompassedwithin the scope of the invention, along with un-solvated and anhydrousforms.

Reference to the compounds of the invention as those of a specificformula or embodiment, e.g., Formula I or any other generic structuralformula or specific compound described or claimed herein, is intended toencompass the specific compound or compounds falling within the scope ofthe formula or embodiment, including salts thereof, particularlypharmaceutically acceptable salts, solvates of such compounds andsolvated salt forms thereof, where such forms are possible unlessspecified otherwise.

Except where noted herein, “alkyl” is intended to include both branched-and straight-chain saturated aliphatic hydrocarbon groups having thespecified number of carbon atoms. Commonly used abbreviations for alkylgroups are used throughout the specification, e.g. methyl may berepresented by conventional abbreviations including “Me” or CH₃ or asymbol that is an extended bond as the terminal group, e.g.,

ethyl may be represented by “Et” or CH₂CH₃, propyl may be represented by“Pr” or CH₂CH₂CH₃, butyl may be represented by “Bu” or CH₂CH₂CH₂CH₃,etc. “C₁₋₄ alkyl” (or “C₁-C₄ alkyl”) for example, means linear orbranched chain alkyl groups, including all isomers, having the specifiednumber of carbon atoms. For example, the structures

have equivalent meanings. C₁₋₄ alkyl includes n-, iso-, sec- andt-butyl, n- and isopropyl, ethyl and methyl. If no number is specified,1-4 carbon atoms are intended for linear or branched alkyl groups.

Also, in the case of a carboxylic acid (—COOH) or alcohol group beingpresent in the compounds of the present invention, pharmaceuticallyacceptable esters of carboxylic acid derivatives, such as methyl, ethyl,or pivaloyloxymethyl, or acyl derivatives of alcohols, such as O-acetyl,O-pivaloyl, O-benzoyl, and O-aminoacyl, can be employed. Included arethose esters and acyl groups known in the art for modifying thesolubility or hydrolysis characteristics for use as sustained-release orprodrug formulations.

If the compounds of Formula I simultaneously contain acidic and basicgroups in the molecule the invention also includes, in addition to thesalt forms mentioned, inner salts or betaines (zwitterions). Salts canbe obtained from the compounds of Formula I by customary methods whichare known to the person skilled in the art, for example by combinationwith an organic or inorganic acid or base in a solvent or dispersant, orby anion exchange or cation exchange from other salts. The presentinvention also includes all salts of the compounds of Formula I which,owing to low physiological compatibility, are not directly suitable foruse in pharmaceuticals but which can be used, for example, asintermediates for chemical reactions or for the preparation ofphysiologically acceptable salts.

Any pharmaceutically acceptable pro-drug modification of a compound ofthe invention which results in conversion in vivo to a compound withinthe scope of the invention is also within the scope of the invention.For example, esters can optionally be made by esterification of anavailable carboxylic acid group or by formation of an ester on anavailable hydroxy group in a compound. Similarly, labile amides can bemade. Pharmaceutically acceptable esters or amides of the compounds ofthe invention may be prepared to act as pro-drugs which can behydrolyzed back to an acid (or —COO— depending on the pH of the fluid ortissue where conversion takes place) or hydroxy form particularly invivo and as such are encompassed within the scope of the invention.Examples of pharmaceutically acceptable pro-drug modifications include,but are not limited to, —C₁₋₆alkyl esters and —C₁₋₆alkyl substitutedwith phenyl esters.

When any variable occurs more than one time in any constituent or informula I, its definition on each occurrence is independent of itsdefinition at every other occurrence. Also, combinations of substituentsand/or variables are permissible only if such combinations result instable compounds.

Except where noted herein, “alkanol” is intended to include aliphaticalcohols having the specified number of carbon atoms, such as methanol,ethanol, propanol, etc., where the —OH group is attached at anyaliphatic carbon, e.g., propan-1-ol, propan-2-ol, etc.

Alkyl groups may be unsubstituted, or substituted with 1 to 3substituents on any one or more carbon atoms, with halogen, C₁-C₂₀alkyl, CF₃, NH₂, —NH(C₁-C₆ alkyl), —N(C₁-C₆ alkyl)₂, NO₂, oxo, CN, N₃,—OH, —O(C₁-C₆ alkyl), C₃-C₁₀ cycloalkyl, C₂-C₆ alkenyl, C2-C₆ alkynyl,(C₁-C₆ alkyl)S(O)₀₋₂—, HS(O)₀₋₂—, (C₁-C₆ alkyl)S(O)₀₋₂(C₁-C₆ alkyl)-,HS(O)₀₋₂(C₁-C₆ alkyl)-, (C₀-C₆ alkyl)C(O)NH—, H₂N—C(NH)—, —O(C₁-C₆alkyl)CF₃, HC(O)—, (C₁-C₆ alkyl)C(O)—, HOC(O)—, (C₁-C₆ alkyl)OC(O)—,HO(C₁-C₆ alkyl)-, (C₁-C₆ alkyl)O(C₁-C₆ alkyl)-, (C₁-C₆alkyl)C(O)₁₋₂(C₁-C₆ alkyl)-, HC(O)₁₋₂(C₁-C₆ alkyl)-, (C₁-C₆alkyl)C(O)₁₋₂—, HOC(O)NH—, (C₁-C₆ alkyl)OC(O)NH—, aryl, aralkyl,heterocycle, heterocyclylalkyl, halo-aryl, halo-aralkyl,halo-heterocycle, halo-heterocyclylalkyl, cyano-aryl, cyano-aralkyl,cyano-heterocycle and cyano-heterocyclylalkyl, where such substitutionresults in formation of a stable compound. Unless otherwise specified,alkyl groups are unsubstituted.

Except where noted, the term “halogen” means fluorine, chlorine, bromineor iodine.

Except where noted, the term “saturated heterocycle” refers to a stable4- to 7-membered mono-cyclic or stable 7- to 12-membered bicyclic orstable 12- to 14-membered tricyclic heteroatom-containing ring systemunsubstituted or substituted with C₁₋₄ alkyl or halogen, and whichconsists of carbon atoms and from one to four heteroatoms independentlyselected from the group consisting of N, O and S, and wherein thenitrogen and sulfur heteroatoms may optionally be oxidized, and thenitrogen heteroatom may optionally be quaternized. Especially useful arerings containing one oxygen or sulfur, one to four nitrogen atoms, orone oxygen or sulfur combined with one or two nitrogen atoms. Theheterocyclic ring may be attached at any heteroatom or carbon atom whichresults in the creation of a stable structure. Representative examplesinclude azetidine, oxetane, thietane, diazetidine, dioxetane,dithietane, pyrrolidine, tetrahydrofuran, thiolane, imidazolidine,pyrazolidine, oxazolidine, isoxazolidine, thiazolidine, isothiazolidine,dioxolane, dithiolane, piperidine, oxane, thiane, piperazine,morpholine, thiomorpholine, dioxane, dithiane, trioxane, trithiane,azepane, oxepane, thiepane and homopiperazine.

Except where noted herein, the term “unsaturated heterocycle” refers toa monocyclic unsaturated heterocycle having a specified number of atommembers (e.g., 4, 5, 6 or 7-membered), including a specified number ofheteroatoms (e.g., 1, 2, 3 or 4 heteroatoms independently selected fromN, O or S), or a bicyclic unsaturated ring system having a specifiednumber of atom members (e.g., 7, 8, 9, 10, 11 or 12-membered) includinga specified number of heteroatoms (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10heteroatoms independently selected from N, S or O) or a tricyclicunsaturated ring system having a specified number of atom members (e.g.,12-, 13- or 14-membered) including a specified number of heteroatoms(e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 heteroatoms independentlyselected from N, S or O) e.g., 5-membered rings containing one nitrogen(pyrrole), one oxygen (furan) or one sulfur (thiophene) atom, 5-memberedrings containing one nitrogen and one sulfur (thiazole) atom, 5-memberedrings containing one nitrogen and one oxygen (oxazole or isoxazole)atom, 5-membered rings containing two nitrogen (imidazole or pyrazole)atoms, five-membered aromatic rings containing three nitrogen (triazole)atoms, five-membered aromatic rings containing one oxygen, one nitrogenor one sulfur atom, five-membered aromatic rings containing twoheteroatoms independently selected from oxygen, nitrogen and sulfur(e.g., oxazole), 6-membered rings containing one nitrogen (pyridine), orone oxygen (pyran) atom, 6-membered rings containing two nitrogen(pyrazine, pyrimidine, or pyridazine) atoms, 6-membered rings containingthree nitrogen (triazine) atoms, a tetrazolyl ring; a thiazinyl ring; orcoumarinyl. Additional examples are pyridine, pyrimidine, thiophene,imidazole, isothiazole, oxadiazole, and isoxazole.

Except where noted herein, the term “unsaturated bicyclic heterocycle”or “unsaturated tricyclic heterocycle” refers to a heterocycle havingfused rings in which at least one of the rings is not fully saturated,e.g.

is a 9-membered unsaturated bicyclic heterocycle having one nitrogenatom.

Except where noted herein, the term “carbocycle” (and variations thereofsuch as “carbocyclic” or “carbocyclyl”) as used herein, unless otherwiseindicated, refers to a C₃ to C₈ monocyclic saturated or unsaturatedring, e.g., C₃₋₈ monocyclic carbocycle, or a C₉ to C₁₂ bicyclicsaturated or unsaturated ring, e.g., C₉₋₁₂ bicyclic carbocycle. Thecarbocycle may be attached to the rest of the molecule at any carbonatom which results in a stable compound. Saturated carbocyclic ringsinclude, for example, “cycloalkyl” rings, e.g., cyclopropyl, cyclobutyl,etc. Unsaturated carbocyclic rings include, for example

Unsaturated bicyclic carbocyclic ring systems include fused ring systemswhere all ring system members are carbon atoms and where at least one ofthe fused rings is not saturated.

Except where noted herein, the term “unsaturated bicyclic carbocycle” or“unsaturated tricyclic carbocycle” refers to a carbocycle having fusedrings in which at least one of the rings is not fully saturated, e.g.

is a 9-membered unsaturated bicyclic carbocycle.

Except where noted, the term “aryl” refers to a stable 6- to 10-memberedmono- or bicyclic unsaturated carbocyclic ring system such as phenyl, ornaphthyl. The aryl ring can be unsubstituted or substituted with one ormore of C₁₋₄ alkyl, hydroxyl, alkoxy, halogen, or amino.

“Celite®” (Fluka) diatomite is diatomaceous earth, and can be referredto as “celite”.

Carbocycle groups may be unsubstituted, or substituted on any one ormore carbon atoms, with halogen, C₁-C₂₀ alkyl, CF₃, NH₂, —NH(C₁-C₆alkyl), —N(C₁-C₆ alkyl)₂, NO₂, oxo, CN, N₃, —OH, —O(C₁-C₆ alkyl), C₃-C₁₀cycloalkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, HS(O)₀₋₂—, (C₁-C₆alkyl)S(O)₀₋₂—, (C₁-C₆ alkyl)S(O)₀₋₂(C₁-C₆ alkyl)-, HS(O)₀₋₂(C₁-C₆alkyl)-, (C₁-C₆ alkyl)S(O)₀₋₂, (C₁-C₆ alkyl)C(O)NH—, HC(O)NH—,H₂N—C(NH)—, —O(C₁-C₆ alkyl)CF₃, (C₁-C₆ alkyl)C(O)—, HC(O)—, (C₁-C₆alkyl)OC(O)—, HOC(O)—, (C₁-C₆ alkyl)O(C₁-C₆ alkyl)-, HO(C₁-C₆ alkyl)-,(C₁-C₆ alkyl)C(O)₁₋₂(C₁-C₆ alkyl)-, (C₁-C₆ alkyl)C(O)₁₋₂—,HC(O)₁₋₂(C₁-C₆ alkyl)-, (C₁-C₆ alkyl)OC(O)NH—, HOC(O)NH—, —P(O)(OH)₂,aryl, aralkyl, heterocycle, heterocyclylalkyl, halo-aryl, halo-aralkyl,halo-heterocycle, halo-heterocyclylalkyl, cyano-aryl, cyano-aralkyl,cyano-heterocycle and cyano-heterocyclylalkyl, where such substitutionresults in formation of a stable compound. Unless otherwise specified,carbocycle groups are unsubstituted.

Heterocycles may be unsubstituted, or substituted on any one or morecarbon atoms, with halogen, C₁-C₂₀ alkyl, CF₃, NH₂, —NH(C₁-C₆ alkyl),—N(C₁-C₆ alkyl)₂, NO₂, oxo, CN, N₃, —OH, —O(C₁-C₆ alkyl), C₃-C₁₀cycloalkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, (C₁-C₆ alkyl)S(O)₀₋₂—,HS(O)₀₋₂—, (C₁-C₆ alkyl)S(O)₀₋₂(C₁-C₆ alkyl)-, HS(O)₀₋₂(C₁-C₆ alkyl)-,(C₁-C₆ alkyl)S(O)₀₋₂—, (C₁-C₆ alkyl)C(O)NH—, HC(O)NH—, H₂N—C(NH)—,—O(C₁-C₆ alkyl)CF₃, HC(O)—, (C₁-C₆ alkyl)C(O)—, (C₁-C₆ alkyl)OC(O)—,HOC(O)—, (C₁-C₆ alkyl)O(C₁-C₆ alkyl)-, HO(C₁-C₆ alkyl)-, (C₁-C₆alkyl)O—, (C₁-C₆ alkyl)C(O)₁₋₂(C₁-C₆ alkyl)-, HC(O)₁₋₂(C₁-C₆ alkyl)-,(C₁-C₆ alkyl)C(O)₁₋₂, (C₁-C₆ alkyl)OC(O)NH—, HOC(O)NH—, silyl groups(including trimethylsilyl, tetramethylsilyl, or supersilyl groups suchas tri(trimethylsilyl)silyl or a silicon group connected to tert butylgroups), aryl, aralkyl, heterocycle, heterocyclylalkyl, halo-aryl,halo-aralkyl, halo-heterocycle, halo-heterocyclylalkyl, cyano-aryl,cyano-aralkyl, cyano-heterocycle or cyano-heterocyclylalkyl.Heterocycles may also be independently substituted with a substituent onany one or more nitrogen atoms, with C₁-C₂₀ alkyl, oxo, C₃-C₁₀cycloalkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, aryl, —C(O)C₁₋₆ alkyl,—C(O)NHC₁-C₆ alkyl, —C(O) NH₂, —C₁-C₆ alkylC(O)NH₂, —C₁-C₆alkylOC(O)NH₂, or independently or additionally substituted with 1substituent on any one or more sulfur atoms, with C₁-C₂₀ alkyl, oxo,C₃-C₁₀ cycloalkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, aryl, where suchsubstitution results in formation of a stable compound. Heterocycles mayalso be substituted as described above on one or more carbon atoms andone or more heteroatoms, where such substitutions result in formation ofa stable compound. Unless otherwise specified, heterocycle groups areunsubstituted.

Except where noted herein, structures containing substituent variablessuch as variable “R” below:

which are depicted as not being attached to any one particular bicyclicring carbon atom, represent structures in which the variable can beoptionally attached to any bicyclic ring carbon atom. For example,variable R shown in the above structure can be attached to any one of 6bicyclic ring carbon atoms i, ii, iii, iv, v or vi.

Except where noted herein, a bicyclic heterocycle can be

a fused bicyclic heterocycle, e.g.,

a bridged bicyclic heterocycle, e.g., or

a spiro bicyclic heterocycle, e.g.

Where ring atoms are represented by variables such as “X”, e.g.,

the variables are defined by indicating the atom located at the variablering position without depicting the ring bonds associated with the atom.For example, when X in the above ring is nitrogen, the definition willshow “N” and will not depict the bonds associated with it, e.g., willnot show “═N—”. Likewise, when X is a carbon atom that is substitutedwith bromide, the definition will show “C—Br” and will not depict thebonds associated with it, e.g., will not show

The invention also includes derivatives of the compound of Formula I,acting as prodrugs and solvates. Prodrugs, following administration tothe patient, are converted in the body by normal metabolic or chemicalprocesses, such as through hydrolysis in the blood, to the compound ofFormula 1. Such prodrugs include those that demonstrate enhancedbioavailability, tissue specificity, and/or cellular delivery, toimprove drug absorption of the compound of Formula I. The effect of suchprodrugs may result from modification of physicochemical properties suchas lipophilicity, molecular weight, charge, and other physicochemicalproperties that determine the permeation properties of the drug.

The preparation of pharmacologically acceptable salts from compounds ofthe Formula (I) capable of salt formation, including theirstereoisomeric forms is carried out in a manner known per se. With basicreagents such as hydroxides, carbonates, hydrogencarbonates, alkoxidesand ammonia or organic bases, for example, trimethyl- or triethylamine,ethanolamine, diethanolamine or triethanolamine, trometamol oralternatively basic amino acids, for example lysine, ornithine orarginine, the compounds of the Formula (I) form stable alkali metal,alkaline earth metal or optionally substituted ammonium salts. If thecompounds of the Formula (I) have basic groups, stable acid additionsalts can also be prepared using strong acids. For the, inorganic andorganic acids such as hydrochloric, hydrobromic, sulfuric, hemisulfuric,phosphoric, methanesulfonic, benzenesulfonic, p-toluenesulfonic,4-bromobenzenesulfonic, cyclohexylamidosulfonic,trifluoromethylsulfonic, 2-hydroxyethanesulfonic, acetic, oxalic,tartaric, succinic, glycerolphosphoric, lactic, malic, adipic, citric,fumaric, maleic, gluconic, glucuronic, palmitic or trifluoroacetic acidare suitable.

The invention also relates to medicaments containing at least onecompound of the Formula (I) and/or of a pharmaceutically acceptable saltof the compound of the Formula (I) and/or an optionally stereoisomericform of the compound of the Formula (I) or a pharmaceutically acceptablesalt of the stereoisomeric form of the compound of Formula (I), togetherwith a pharmaceutically suitable and pharmaceutically acceptablevehicle, additive and/or other active substances and auxiliaries.

The medicaments according to the invention can be administered by oral,inhalative, rectal or transdermal administration or by subcutaneous,intraarticular, intraperitoneal or intravenous injection. Oraladministration is preferred. Coating of stents with compounds of theFormula (I) and other surfaces which come into contact with blood in thebody is possible.

The invention also relates to a process for the production of amedicament, which comprises bringing at least one compound of theFormula (I) into a suitable administration form using a pharmaceuticallysuitable and pharmaceutically acceptable carrier and optionally furthersuitable active substances, additives or auxiliaries.

Suitable solid or galenical preparation forms are, for example,granules, powders, coated tablets, tablets, (micro)capsules,suppositories, syrups, juices, suspensions, emulsions, drops orinjectable solutions and preparations having prolonged release of activesubstance, in whose preparation customary excipients such as vehicles,disintegrants, binders, coating agents, swelling agents, glidants orlubricants, flavorings, sweeteners and solubilizers are used. Frequentlyused auxiliaries which may be mentioned are magnesium carbonate,titanium dioxide, lactose, mannitol and other sugars, talc, lactose,gelatin, starch, cellulose and its derivatives, animal and plant oilssuch as cod liver oil, sunflower, peanut or sesame oil, polyethyleneglycol and solvents such as, for example, sterile water and mono- orpolyhydric alcohols such as glycerol.

The dosage regimen utilizing the compounds is selected in accordancewith a variety of factors including type, species, age, weight, sex andmedical condition of the patient; the severity of the condition to betreated; the route of administration; the renal and hepatic function ofthe patient; and the particular compound or salt thereof employed. Anordinarily skilled physician or veterinarian can readily determine andprescribe the effective amount of the drug required to prevent, counter,or arrest the progress of the condition.

Oral dosages of the compounds, when used for the indicated effects, willrange between about 0.01 mg per kg of body weight per day (mg/kg/day) toabout 30 mg/kg/day, preferably 0.025-7.5 mg/kg/day, more preferably0.1-2.5 mg/kg/day, and most preferably 0.1-0.5 mg/kg/day (unlessspecified otherwise, amounts of active ingredients are on free basebasis). For example, an 80 kg patient would receive between about 0.8mg/day and 2.4 g/day, preferably 2-600 mg/day, more preferably 8-200mg/day, and most preferably 8-40 mg/kg/day. A suitably preparedmedicament for once a day administration would thus contain between 0.8mg and 2.4 g, preferably between 2 mg and 600 mg, more preferablybetween 8 mg and 200 mg, and most preferably 8 mg and 40 mg, e.g., 8 mg,10 mg, 20 mg and 40 mg. Advantageously, the compounds may beadministered in divided doses of two, three, or four times daily. Foradministration twice a day, a suitably prepared medicament would containbetween 0.4 mg and 4 g, preferably between 1 mg and 300 mg, morepreferably between 4 mg and 100 mg, and most preferably 4 mg and 20 mg,e.g., 4 mg, 5 mg, 10 mg and 20 mg.

Intravenously, the patient would receive the active ingredient inquantities sufficient to deliver about 0.01 mg per kg of body weight perday (mg/kg/day) to about 30 mg/kg/day, preferably 0.025-7.5 mg/kg/day,more preferably 0.1-2.5 mg/kg/day, and even more preferably 0.1-0.5mg/kg/day. Such quantities may be administered in a number of suitableways, e.g. large volumes of low concentrations of active ingredientduring one extended period of time or several times a day, low volumesof high concentrations of active ingredient during a short period oftime, e.g. once a day. Typically, a conventional intravenous formulationmay be prepared which contains a concentration of active ingredient ofbetween about 0.01-1.0 mg/ml, e.g. 0.1 mg/ml, 0.3 mg/ml, and 0.6 mg/ml,and administered in amounts per day of between 0.01 ml/kg patient weightand 10.0 ml/kg patient weight, e.g. 0.1 ml/kg, 0.2 ml/kg, 0.5 ml/kg. Inone example, an 80 kg patient, receiving 8 ml twice a day of anintravenous formulation having a concentration of active ingredient of0.5 mg/ml, receives 8 mg of active ingredient per day. Glucuronic acid,L-lactic acid, acetic acid, citric acid or any pharmaceuticallyacceptable acid/conjugate base with reasonable buffering capacity in thepH range acceptable for intravenous administration may be used asbuffers. The choice of appropriate buffer and pH of a formulation,depending on solubility of the drug to be administered, is readily madeby a person having ordinary skill in the art.

The compounds of the invention may be prepared by employing reactions asshown in the following Reaction Schemes, in addition to other standardmanipulations that are known in the literature or exemplified in theexperimental procedures. The illustrative Reaction Schemes below,therefore, are not limited by the compounds listed or by any particularsubstituents employed for illustrative purposes. Substituent numberingas shown in the Reaction Schemes do not necessarily correlate to thatused in the claims and often, for clarity, a single substituent is shownattached to the compound where multiple substituents are optionallyallowed under the definitions of Formula I hereinabove.

Methods for Making the Compounds of Present Invention General Methods

The compounds of the present invention can be readily produced fromknown compounds or commercially available compounds by, for example,known processes described in published documents, and produced byproduction processes described below. The present invention is notlimited to the production processes described below. The invention alsoincludes processes for the preparation of compounds of the invention.

It should be noted that, when a compound of structural Formula I has areactive group such as hydroxy group, amino group, carboxyl group, orthiol group as its substituent, such group may be adequately protectedwith a protective group in each reaction step and the protective groupmay be removed at an adequate stage. The process of such introductionand removal of the protective group may be adequately determineddepending on the group to be protected and the type of the protectivegroup, and such introduction and removal are conducted, for example, bythe process described in the review section of Greene, T. W., et. al.,“Protective Groups in Organic Synthesis”, 2007, 4th Ed., Wiley, NewYork, or Kocienski, P., “Protecting Groups” 1994, Thieme.

The present invention is not limited in scope by the specificembodiments disclosed in the examples which are intended asillustrations of a few aspects of the invention and any embodiments thatare functionally equivalent are within the scope of this invention.Indeed, various modifications of the invention in addition to thoseshown and described herein will become apparent to those skilled in therelevant art and are intended to fall within the scope of the appendedclaim.

All solvents used were commercially available and were used withoutfurther purification. Reactions were typically run using anhydroussolvents under an inert atmosphere of nitrogen.

¹H spectra were recorded at 300 or 400 MHz for proton on a BrukerMercury Plus 400 NMR Spectrometer equipped with a Bruker 400 BBO probe.All deuterated solvents contained typically 0.03% to 0.05% v/vtetramethylsilane, which was used as the reference signal (set at 6 0.00for both ¹H and ¹³C).

LCMS analyses were performed on a SHIMADZU LCMS consisting of an UFLC20-AD and LCMS 2020 MS detector. The column used was a Shim-pack XR-ODS,2.2 μm, 3.0×50 mm. A linear gradient was applied, starting at 95% A (A:0.05% TFA in water) and ending at 100% B (B: 0.05% TFA in MeCN) over 2.2min with a total run time of 3.6 min. The column temperature was at 40°C. with the flow rate of 1.0 mL/min. The Diode Array Detector wasscanned from 200-400 nm. The mass spectrometer was equipped with anelectrospray ion source (ES) operated in a positive or negative mode.

HPLC analyses were performed on a SHIMADZU UFLC with two LC20 AD pumpand a SPD-M20A Photodiode Array Detector. The column used was an XBridgeC₁₈, 3.5 μm, 4.6×100 mm. A linear gradient was applied, starting at 90%A (A: 0.05% TFA in water) and ending at 95% B (B: 0.05% TFA in MeCN)over 10 min with a total run time of 15 min. The column temperature wasat 40° C. with the flow rate of 1.5 mL/min. The Diode Array Detector wasscanned from 200-400 nm.

Thin layer chromatography (TLC) was performed on Alugram® (Silica gel 60F₂₅₄) from Mancherey-Nagel and UV was typically used to visualize thespots. Additional visualization methods were also employed in somecases. In these cases the TLC plate was developed with iodine (generatedby adding approximately 1 g of I₂ to 10 g silica gel and thoroughlymixing), ninhydrin (available commercially from Aldrich), or Magic Stain(generated by thoroughly mixing 25 g (NH₄)₆Mo₇O₂₄.4H₂, 5 g(NH₄)₂Ce(IV)(NO₃)₆ in 450 mL water and 50 mL concentrated H₂SO₄) tovisualize the compound. Flash chromatography was preformed using 40-63μm (230-400 mesh) silica gel from Silicycle following analogoustechniques to those disclosed in Still, W. C.; Kahn, M.; and Mitra, M.Journal of Organic Chemistry, 1978, 43, 2923. Typical solvents used forflash chromatography or thin layer chromatography were mixtures ofchloroform/methanol, dichloromethane/methanol, ethyl acetate/methanoland petroleum ether/ethyl acetate.

Preparative HPLC was performed on either a Waters Prep LC 4000 Systemusing a Waters 2487 Diode Array or on a Waters LC Module 1 plus. Thecolumn used was SunFire Prep C18 OBD Column, 5 m, 19×150 mm. Narrowgradients with acetonitrile/water, with the water containing either 0.1%trifluoroacetic acid or 0.1% NH₄HCO₃, were used to elute the compound ata flow rate of 20 mL/min and a total run time between 20-30 min.Detector, 254 nm, 220 nm.

Chiral HPLC conditions: Column, Chiralpak IA, 5 m, 20×150 mm; Mobilephase, Hex/EtOH or IPA; Detector, 254 nm, 220 nm.

Starting materials used were either available from commercial sources orprepared according to literature procedures and had experimental data inaccordance with those reported.

The following abbreviations have been used:

-   ACN acetonitrile-   AcOH acetic acid-   (Boc)₂O di-tert-butyl dicarbonate-   BPO benzoyl peroxide-   BSA bovine serum albumin-   ° C. degree Celsius-   CCl₄ carbon tetrachloride-   CDCl₃ deuterated chloroform-   CD₃OD deuterated methanol-   CO carbon monoxide-   DCM dichloromethane-   DIEA N,N-disisopropylethylamine-   DMA dimethylacetamide-   DMAP 4-(dimethylamino)pyridine-   DMF N,N-dimethylformamide-   DMS dimethyl sulfide-   DMSO dimethylsulfoxide-   DTT dithiothreitol-   EDC N-(3-dimethylaminpopropyl)-N′-ethylcarbodiimide-   EtOAc ethyl acetate-   EtOH ethanol-   g gram-   h hour(s)-   HATU    N-[(Dimethylamino)-1H-1,2,3-triazolo-[4,5-b]pyridin-1-ylmethylene]-N-methylmethanaminium    hexafluorophosphate N-oxide-   HCl hydrochloric acid-   HPLC high pressure liquid chromatography-   LCMS liquid chromatography and mass spectrometry-   M molar-   MeOH methanol-   μL microliter-   MS mass spectrometry-   MTBE methyl tert-butyl ether-   mmol millimole-   mg milligram-   min minutes-   mL milliliter(s)-   nM nanomolar-   N normal-   NaHMDS sodiumbis(trimethylsilyl)amide-   Na₂SO₄ sodium sulfate-   NBS N-bromosuccinimide-   NH₄HCO₃ ammonium bicarbonate-   NMR nuclear magnetic resonance-   Pd(dtbpf)Cl₂    [1,1′-Bis(di-tert-butylphosphino)ferrocene]dichloropalladium(II)-   PdCl₂(dppf)    [1,1-bis(diphenylphosphine)ferrocene]dichloropalladium(II)-   Pd(PPh₃)₄ tetrakis(triphenylphosphine)palladium(O)-   pet. ether petroleum ether-   POCl₃ phosphorus(V) oxychloride-   Prep-TLC preparative TLC-   psi pound per square inch-   RuPhos Pd G₁    Chloro(2-dicyclohexylphosphino-2′,6′-di-i-propoxy-1,1′-biphenyl)[2-(2-aminoethylphenyl)]palladium(II),    methyl-t-butylether adduct-   Schwartz's reagent zirconocene hydrochloride-   SFC supercritical fluid chromatography-   SOCl₂ thionyl chloride-   T3P Propylphosphonic anhydride-   TEA triethylamine-   Tf trifluoromethanesulfonate-   TFA trifluoroaceticacid-   THE tetrahydrofuran-   TLC thin layer chromatography-   Xphos-Pd-G3    (2-Dicyclohexylphosphino-2′,4′,6′-triisopropyl-1,1′-biphenyl)[2-(2′-amino-1,1′-biphenyl)]palladium(II)    methanesulfonate-   XantPhos-Pd-G2    Chloro[(4,5-bis(diphenylphosphino)-9,9-dimethylxanthene)-2-(2′-amino-1,1′-biphenyl)]palladium(II)

General Synthetic Schemes

While the present invention has been described in conjunction with thespecific examples set forth above, many alternatives, modifications andvariations thereof will be apparent to those of ordinary skill in theart. All such alternatives, modifications and variations are intended tofall within the spirit and scope of the present invention.

Triazole 3 can be formed via a two-step sequence. Reaction of hydrazine1 with formamide gives triazole 2 which undergoes bromination to givetriazole 3.

Compound 9 can be formed in the following sequence. Alkylation of acid 4affords ester 5, while LiBH₄ reduction followed by Swern oxidationaffords aldehyde 7. Alkyl magnesium chloride addition to aldehyde 7affords alcohol 8, which cyclizes under basic conditions to give epoxide9.

Copper catalyst 11 can be formed in one step from diamine 10 andCu(OAc)₂.

Compound 13 can be formed via a two-step sequence. Copper catalyzednitration of aldehyde 7 followed by nitro reduction affords aminoalcohol 13.

Compound 15 can be formed by alkylation of 14 under basic conditions.

Compound 16 can be formed by dimethylation 14 under basic conditions.

Compound of formula 21 can be formed in the following sequence.Reduction of nitrile 17 affords amine 18, which can be converted tocarbamate 20 with carbonate 19 under basic conditions. Acid mediatedcyclization affords lactam 21.

Compound of formula 22 can be formed by epoxide ring opening of 9 withlactam 21. Compound 22 can be used to prepare a number of intermediatesas shown in the scheme below:

Deprotection under basic conditions affords tetrahydroisoquinolinone 23and boc protection affords tetrahydroisoquinolinone 24, which can beconverted back to 23 under acidic conditions. A Pd-catalyzed C—Hinsertion between aryl bromide 22 and triazole 25 affords 26, which uponbasic deprotection affords tetrahydroisoquinolinone 27. Carbonylation ofaryl bromide 22 with CO in EtOH affords ester 28, followed by hydrolysisto give acid 29. Amide coupling between acid 29 and bridged morpholine30 affords amide 31. Deprotection under basic conditions affordstetrahydroisoquinolinone 32.

Compound 39 can be formed in the following sequence. Esterification ofacid 33 affords ester 34. Benzylic bromination of 34 affordsbenzylbromide 35, which undergoes cyanation to give benzyl cyanide 36.Double alkylation at benzylic position affords cyclobutane 37. Treatmentof cyanide 37 with Schwartz's reagent affords aldehyde 38, whichundergoes reductive amination and cyclization to afford lactam 39.

Compound of formula 43 can be formed in the following sequence.Pd-catalyzed borylation of aryl bromide 40 affords a mixture of boronicacid and boronic ester 41. A Suzuki reaction with triazole 3 affordstetrahydroisoquinolinone 42. Deprotection under acidic conditionsaffords tetrahydroisoquinolinone 43.

Compound of formula 47 can be formed in the following sequence.Carbonylation of aryl bromide 21 with CO in EtOH affords ester 44,followed by hydrolysis to give acid 45. Amide coupling between acid 29and bridged morpholine affords amide 46. Reacting lactam 46 with epoxide9 affords ring opening product 47.

Compounds of formula 5 can be formed in the following sequence. Reactinglactam 21 with epoxide 9 affords compound 48. Carbonylation of arylbromide 48 affords ester 49, which upon hydrolysis affords acid 50.Amide coupling between acid 50 and amines affords 51. Final deprotectionunder acidic conditions affords tetrahydroisoquinolinone 52.

Compound 55 can be formed via a two-step sequence. Pd-catalyzed C—Ncross coupling between aryl bromide 23 and lactam 53 affordstetrahydroisoquinolinone 54. Deprotection under acidic conditionsaffords tetrahydroisoquinolinone 55.

INTERMEDIATES Intermediate 1: tert-butyl(3S)-3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate

Step 1:

To a mixture of(S)-2-(tert-butoxycarbonyl)-1,2,3,4-tetrahydroisoquinoline-3-carboxylicacid (100 g, 361 mmol) and K₂CO₃ (150 g, 1082 mmol) in MeOH (500 ml) wasadded a solution of MeI (120 ml, 1916 mmol) in DMF (150 ml) at 30° C.The resulting solution was stirred at 30° C. for 72 h. Water (1500 mL)was then added and the mixture was extracted with EtOAc (500 mL×3). Theseparated organic layers were concentrated in vacuum and the resultingresidue was purified by flash silica gel chromatography (eluting withethyl acetate/pet. ether gradient) to give (S)-2-tert-butyl 3-methyl3,4-dihydroisoquinoline-2,3(1H)-dicarboxylate as colorless oil. MS: 314(M+23).

Step 2:

To a solution of (S)-2-tert-butyl 3-methyl3,4-dihydroisoquinoline-2,3(1H)-dicarboxylate (105 g, 360 mmol) in THF(600 ml) was added LiBH₄ (20.41 g, 937 mmol) slowly at 30° C. Themixture was stirred at 30° C. for 12 h. The mixture was diluted withwater (1500 mL) and with ethyl acetate (200 mL). The resulting mixturewas extracted with ethyl acetate (100 mL×3). The combined organic layerswere washed with brine, dried over anhydrous Na₂SO₄, filtered, andconcentrated. The residue was purified by flash silica gelchromatography (eluting with ethyl acetate/pet. ether gradient) to give(S)-tert-butyl3-(hydroxymethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate as colorlessoil. MS: 286 (M+23).

Step 3:

To a solution of oxalyl dichloride (32.4 ml, 383 mmol) in DCM (1300 mL)was added DMSO (54.3 ml, 766 mmol) in DCM (200 mL) at −78° C. Themixture was stirred at −78° C. for 30 min. To the mixture was then added(S)-tert-butyl3-(hydroxymethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (56 g, 213mmol) in DCM (500 mL) at −78° C. The resulting mixture was stirred at−78° C. for 30 min. To the mixture was added TEA (213 ml, 1531 mmol) at−78° C. The resulting mixture was stirred at −78° C. for 30 min. Themixture was then diluted with water (1600 mL) and extracted with DCM(500 mL). The organic layer was adjusted to pH-5 with aqueous HCl (0.1M). The resulting mixture was washed with brine (1000 mL). The organiclayer was dried over anhydrous Na₂SO₄, filtered, and concentrated togive the crude product (S)-tert-butyl3-formyl-3,4-dihydroisoquinoline-2(1H)-carboxylate as yellow oil. Thismaterial was used without further purification in the next step.

Step 4:

To a stirred solution of chloroiodomethane (46.6 g, 264 mmol) in THE(1500 ml) was added isopropylmagnesium chloride (106 ml, 211 mmol) at−78° C. under N₂ atmosphere. The solution was stirred at −78° C. for 20min and then (S)-tert-butyl3-formyl-3,4-dihydroisoquinoline-2(1H)-carboxylate (46 g, 176 mmol) inTHE (300 ml) was added. The resulting solution was stirred for 2 h at−78° C. The mixture was then diluted with saturated aqueous NH₄Cl (1000mL) and EtOAc (500 mL). The mixture was extracted with EtOAc (500 mL×3).The combined organic layers were washed with brine (400 mL), dried overanhydrous Na₂SO₄, filtered, and concentrated. The residue was purifiedby flash silica gel chromatography (eluting with ethyl acetate/pet.ether gradient) to give (3S)-tert-butyl3-(2-chloro-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate asyellow oil. MS: 334 (M+23).

Step 5:

To a stirred solution of (3S)-tert-butyl3-(2-chloro-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(15 g, 48.1 mmol) in THE (75 ml) was added KOH (13.17 g, 192 mmol) inMeOH (50 ml) dropwise at 0° C. The resulting solution was stirred at 20°C. for 0.5 h. The mixture was diluted with water (250 mL) and EtOAc (100mL). The resulting mixture was extracted with EtOAc (100 mL×3). Thecombined organic layers were washed with brine (50 mL), dried overanhydrous Na₂SO₄, filtered, and concentrated to give (3S)-tert-butyl3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate as brown oilwhich was used directly in the next step. MS: 298 (M+23).

Intermediate 2:tert-butyl(S)-3-((R)-2-amino-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate

Step 1:

Copper diamine catalyst synthesis: To a mixture of(1R,2R)—N¹,N²-bis(4-chlorobenzyl)cyclohexane-1,2-diamine (7 g, 19.27mmol) in EtOH (200 ml) at 30° C. was added copper(II) acetatemonohydrate (3.85 g, 19.27 mmol). The reaction was stirred for 12 h. Theresidue was concentrated under reduced pressure to remove residual EtOH.The foam was purified by flash silica gel chromatography (EtOH/DCM aseluant) to give copper diamine catalyst as a blue solid which was usedwithout further purification.

Step 2:

To a solution of (S)-tert-butyl3-formyl-3,4-dihydroisoquinoline-2(1H)-carboxylate (6 g, 22.96 mmol) inethanol (40 ml) was added the copper diamine catalyst prepared in step 1(2.49 g, 4.59 mmol) in ethanol (10 ml) at 30° C. The solution was cooledto 0° C., nitromethane (14.02 g, 230 mmol) was added, and the reactionwas stirred at 30° C. for 2 days. The mixture was diluted with water andextracted with EtOAc. The combined organic layers were washed withbrine, dried over anhydrous Na₂SO₄, filtered, and concentrated. Theresidue was purified by flash silica gel chromatography (eluting withethyl acetate/pet. ether gradient) to give (S)-tert-butyl3-((R)-1-hydroxy-2-nitroethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas yellow oil. The product was further purified byprep-HPLC(acetonitrile/water+0.1% TFA modifier) to give (S)-tert-butyl3-((R)-1-hydroxy-2-nitroethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(3.1 g) as a yellow oil. MS: 267 (M−t-Bu+H).

Step 3:

To a solution of (S)-tert-butyl3-((R)-1-hydroxy-2-nitroethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(3.1 g, 9.62 mmol) in MeOH (20 ml) was added Raney nickel (2 g, 34.1mmol) under N₂ atmosphere. The mixture was degassed and backfilled withH₂ (three times). The resulting mixture was stirred under 15 psi of H₂at 29° C. for 2 h. The catalyst was filtered off and filtrate wasconcentrated under reduced pressure to give (S)-tert-butyl3-((R)-2-amino-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas yellow oil, which was used directly in the next step. MS: 293 (M+1).

Intermediate 3: 5-bromo-1-cyclopropyl-1H-1,2,4-triazole

Step 1:

A solution of cyclopropylhydrazine hydrochloride (483 mg, 4.45 mmol) informamide (3.5 mL, 89 mmol) was heated at 130° C. for 3 days. Thesolution was cooled to room temperature, diluted with brine, andextracted with EtOAc (3×). The combined organic layers were dried oversodium sulfate, filtered, and concentrated under reduced pressure(material is volatile). The residue was purified by columnchromatography on silica (0-15% MeOH/DCM) to afford1-cyclopropyl-1H-1,2,4-triazole as a liquid. ¹H NMR (500 MHz, CDCl₃) δ:8.22 (s, 1H), 7.93 (s, 1H), 3.67-3.58 (m, 1H), 1.23-1.07 (m, 4H).

Step 2:

To a −78° C. solution of 1-cyclopropyl-1H-1,2,4-triazole (403 mg, 3.69mmol) in THE (25 mL) under an atmosphere of nitrogen was added nBuLi(2.5 M in hexanes, 1.6 mL, 4.1 mmol). The solution was stirred for 45min, and then 1,2-dibromotetrafluoroethane (0.48 mL, 4.1 mmol) wasadded. The solution was warmed to room temperature and stirred for 2 h.The mixture was diluted with EtOAc, washed with water and then brine,dried over sodium sulfate, filtered, and concentrated under reducedpressure. The residue was purified by column chromatography on silica(eluting with MeOH/DCM) to afford5-bromo-1-cyclopropyl-1H-1,2,4-triazole as a solid. MS: 188/190(M+1/M+3). ¹H NMR (500 MHz, CDCl₃) δ: 7.81 (s, 1H), 3.55-3.47 (m, 1H),1.28-1.12 (m, 4H).

Intermediates 4-7: Tert-butyl(3S)-3-[2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl]-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 1 (Intermediate 4), Tert-butyl(3S)-3-[2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl]-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 2 (Intermediate 5), Tert-butyl(3S)-3-[2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl]-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 3 (Intermediate 6), and Tert-butyl(3S)-3-[2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl]-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 4 (Intermediate 7)

Step 1:

To a mixture of 2-(3-bromophenyl)acetonitrile (60 g, 306 mmol) in THE(800 mL) was slowly added NaHMDS (337 mL, 337 mmol) at −78° C. Themixture was stirred at −78° C. for 0.5 h. Then to the mixture was addediodoethane (24.61 mL, 306 mmol) at −78° C. and the reaction was kept at−78° C. for 2 h. The reaction was quenched with saturated aqueous NH₄Cl(200 mL). Then water (200 mL) and EtOAc (300 mL) were added. The organiclayer was separated and the aqueous was re-extracted with EtOAc (300mL×3). The combined organic layers were washed with brine (200 mL),dried over anhydrous Na₂SO₄, filtered, and concentrated. The residue waspurified by flash silica gel chromatography (eluting with ethylacetate/pet. ether gradient) to give 2-(3-bromophenyl)butanenitrile aslight yellow oil. ¹H NMR (400 MHz, CDCl₃) δ 7.42-7.50 (m, 2H), 7.23-7.29(m, 2H), 3.70 (t, J=7.17 Hz, 1H), 1.87-1.97 (m, 2H), 1.06 (t, J=7.28 Hz,3H).

Step 2:

To a mixture of 2-(3-bromophenyl)butanenitrile (56 g, 250 mmol) in THE(1000 mL) was added borane dimethylsulfide complex (75.0 ml, 750 mmol)at 0° C. The resulting mixture was stirred at 30° C. for 12 h. Thereaction was quenched with MeOH (200 mL) at 0° C. Then the mixture wasstirred at 60° C. for 1 h. The mixture was concentrated under reducedpressure to give 2-(3-bromophenyl)butan-1-amine as colourless oil whichwas used in the next step directly. MS: 228/230 (M+1/M+3). ¹H NMR (400MHz, CDCl₃) δ 7.28-7.37 (m, 2H), 7.17 (br t, J=7.39 Hz, 1H), 7.05-7.12(m, 1H), 2.87-2.96 (m, 1H), 2.76-2.87 (m, 1H), 2.37-2.54 (m, 1H),1.65-1.69 (m, 1H), 1.44-1.57 (m, 1H), 0.74-0.85 (m, 3H).

Step 3:

To a mixture of 2-(3-bromophenyl)butan-1-amine (132 g, 579 mmol) and TEA(161 mL, 1157 mmol) in DCM (1500 mL) was added 2,5-dioxopyrrolidin-1-ylmethyl carbonate (120 g, 694 mmol) at 28° C. The mixture was stirred at28° C. for 1 h. The mixture was concentrated under reduced pressure. Tothe mixture was added saturated aqueous NaHCO₃ (400 mL) and H₂O (400mL). The aqueous was re-extracted with EtOAc (700 mL×3). The combinedorganic layers were washed with brine (500 mL), dried over anhydrousNa₂SO₄, filtered and concentrated. The residue was purified by flashsilica gel chromatography (eluting with ethyl acetate/pet. ethergradient) to give methyl (2-(3-bromophenyl)butyl)carbamate as colourlessoil. MS: 286/288 (M+1/M+3). H NMR (400 MHz, CDCl₃) δ 7.35-7.39 (m, 1H),7.30 (s, 1H), 7.19 (t, J=7.63 Hz, 1H), 7.09 (br d, J=7.43 Hz, 1H), 4.51(br s, 1H), 3.62 (s, 3H), 3.53-3.62 (m, 1H), 3.14-3.21 (m, 1H), 2.64 (brs, 1H), 1.63-1.77 (m, 1H), 1.48-1.56 (m, 1H), 0.81 (t, J=7.24 Hz, 3H).

Step 4:

A mixture of methyl (2-(3-bromophenyl)butyl)carbamate (30 g, 105 mmol)and trifluoromethanesulfonic acid (464 mL, 5242 mmol) was stirred at 75°C. for 12 h. The mixture was slowly added into ice water (4 L) andextracted with DCM (3 L x 3). The combined organic layers were washedwith brine (2 L), dried over anhydrous Na₂SO₄, filtered, andconcentrated. The residue was purified by Prep-HPLC (ACN/water with 0.1%TFA modifier) to give 6-bromo-4-ethyl-3,4-dihydroisoquinolin-1(2H)-oneas a white solid. MS: 254/256 (M+1/M+3). ¹H NMR (400 MHz, CDCl₃) δ 7.89(d, J=8.38 Hz, 1H), 7.50 (dd, J=1.76, 8.38 Hz, 1H), 7.39 (d, J=1.76 Hz,1H), 3.64-3.75 (m, 1H), 3.41-3.45 (m, 1H), 2.74 (br dd, J=3.53, 6.84 Hz,1H), 1.65-1.76 (m, 2H), 0.94-1.00 (m, 3H).

Step 5:

To a solution of NaH (0.708 g, 17.71 mmol) in DMF (50 mL) was added6-bromo-4-ethyl-3,4-dihydroisoquinolin-1(2H)-one (3 g, 11.81 mmol). Thereaction mixture was allowed to stir at 20° C. for 0.5 h. Then(3S)-tert-butyl3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (3.25 g, 11.81mmol) was added to the mixture. The reaction mixture was stirred at 40°C. for 12 h. The mixture was slowly added into water (2 L) and extractedwith EtOAc (1.5 L×3). The combined organic layers were washed with brine(2 L), dried over anhydrous Na₂SO₄, filtered and concentrated. Theresidue was purified by Prep-HPLC (ACN/water with 0.1% TFA modifier) togive(10aS)-1-((6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneas brown oil. MS: 455/457 (M+1/M+3). ¹H NMR (400 MHz, CDCl₃) δ 7.86-7.95(m, 1H), 7.53 (ddd, J=1.98, 5.17, 8.25 Hz, 1H), 7.38-7.43 (m, 1H),7.22-7.28 (m, 2H), 7.10-7.22 (m, 2H), 4.60-4.99 (m, 2H), 4.33-4.53 (m,1H), 4.03-4.12 (m, 3H), 3.61-3.91 (m, 2H), 3.09-3.29 (m, 1H), 2.88-3.01(m, 1H), 2.79 (br d, J=3.96 Hz, 1H), 1.63-1.87 (m, 2H), 0.94-1.09 (m,3H).

Step 6:

To a mixture of(10aS)-1-((6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-one(4 g, 8.78 mmol) in MeOH (20 mL) and water (20 mL) was added NaOH (0.703g, 17.57 mmol) at 20° C. The solution was stirred at 80° C. for 36 h.The mixture was concentrated under reduced pressure to give6-bromo-4-ethyl-2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-3,4-dihydroisoquinolin-1(2H)-oneas a light brown solid which was used in next step without purification.MS: 429/431 (M+1/M+3).

Step 7:

To a solution of6-bromo-4-ethyl-2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-3,4-dihydroisoquinolin-1(2H)-one(7.3 g, 17.0 mmol) and TEA (4.74 mL, 34.0 mmol) in DCM (100 mL) wasadded di-tert-butyl dicarbonate (5.92 mL, 25.5 mmol). The mixture wasstirred at 20° C. for 0.5 h. The mixture was then concentrated underreduced pressure. The residue was purified by flash silica gelchromatography (eluting with ethyl acetate/pet. ether gradient) to give(3S)-tert-butyl3-(2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas light yellow oil. MS: 551/553 (M+23/M+25). ¹H NMR (400 MHz, CDCl₃) δ7.83-7.93 (m, 1H), 7.42-7.49 (m, 1H), 7.32 (d, J=3.01 Hz, 1H), 7.19 (brd, J=2.01 Hz, 3H), 7.12 (br s, 1H), 4.60-5.05 (m, 1H), 4.21-4.50 (m,2H), 3.66-3.95 (m, 3H), 3.49 (s, 2H), 2.87-3.32 (m, 2H), 2.68 (br d,J=7.53 Hz, 1H), 1.66 (br d, J=7.53 Hz, 2H), 1.43-1.56 (m, 9H), 0.85-1.04(m, 3H).

Step 8:

The mixture of four stereoisomers of (3S)-tert-butyl3-(2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylatewas separated by chiral SFC (AD column, 40%/60% IPA/CO₂) to afford:

Intermediate 4: (3S)-tert-butyl3-(2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 1 (first eluting): ¹H NMR (400 MHz, CD3OD) δ 7.74 (br d, J=7.89Hz, 1H), 7.42-7.54 (m, 2H), 7.08-7.24 (m, 4H), 4.24-4.60 (m, 2H),4.02-4.21 (m, 1H), 3.91-4.00 (m, 1H), 3.56-3.89 (m, 1H), 3.45-3.49 (m,1H), 3.34 (s, 1H), 3.00-3.25 (m, 2H), 2.90-2.94 (m, 1H), 2.77 (br s,1H), 1.55-1.71 (m, 2H), 1.51 (s, 9H), 0.94 (t, J=7.24 Hz, 3H)

Intermediate 5: (3S)-tert-butyl3-(2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 2 (second eluting): ¹H NMR (400 MHz, CD₃OD) δ 7.67-7.82 (m, 1H),7.39-7.54 (m, 2H), 7.16 (br s, 4H), 4.24-4.62 (m, 2H), 3.90-4.08 (m,1H), 3.73-3.87 (m, 2H), 3.65-3.68 (m, 1H), 3.18-3.22 (m, 1H), 3.00-3.11(m, 1H), 2.90-2.95 (m, 1H), 2.77 (br s, 1H), 1.56-1.75 (m, 2H), 1.51 (s,9H), 0.88-1.02 (m, 3H).

Intermediate 6: (3S)-tert-butyl3-(2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 3 (third eluting): ¹H NMR (400 MHz, CD₃OD) δ 7.80 (br s, 1H),7.41-7.57 (m, 2H), 7.06-7.23 (m, 4H), 4.77-4.82 (m, 1H), 4.27-4.47 (m,2H), 3.78 (br s, 2H), 3.53-3.68 (m, 2H), 3.34 (s, 1H), 3.04 (br s, 2H),2.78 (br s, 1H), 1.55-1.71 (m, 2H), 1.34-1.54 (m, 9H), 0.94 (t, J=7.45Hz, 3H)

Intermediate 7: (3S)-tert-butyl3-(2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 4 (fourth eluting): ¹H NMR (400 MHz, CD₃OD) δ 7.80 (br s, 1H),7.52 (br d, J=8.36 Hz, 1H), 7.48 (s, 1H), 7.07-7.26 (m, 4H), 4.81 (br s,1H), 4.24-4.49 (m, 2H), 3.77-3.94 (m, 3H), 3.52-3.56 (m, 1H), 3.36-3.48(m, 1H), 2.96-3.16 (m, 2H), 2.77 (br s, 1H), 1.59-1.75 (m, 2H),1.42-1.56 (m, 9H), 0.90-1.01 (m, 3H).

Intermediate 8 and 9: (S)-tert-butyl3-((R)-2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(Intermediate 8) and (S)-tert-butyl3-((S)-2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(Intermediate 9)

Step 1:

To a mixture of 2-(3-bromophenyl)acetonitrile (30 g, 153 mmol) in THE(600 mL) was slowly added NaHMDS (383 mL, 383 mmol) at −78° C. Themixture was stirred at −78° C. for 0.5 h. Then to the mixture was addediodomethane (31.3 mL, 418 mmol) at −78° C. The mixture was stirred at−78° C. for 2 h. The reaction was then quenched with saturated aqueousNH₄Cl (200 mL). Water (200 mL) and EtOAc (300 mL) were added. Theorganic layer was separated and the aqueous layer was re-extracted withEtOAc (300 mL×3). The combined organic layers were washed with brine(500 mL), dried over anhydrous Na₂SO₄, and concentrated. The residue waspurified by flash silica gel chromatography (eluting with ethylacetate/pet. ether gradient) to give2-(3-bromophenyl)-2-methylpropanenitrile (37 g) as brown oil. ¹H NMR(400 MHz, CDCl₃) δ 7.58 (t, J=1.87 Hz, 1H), 7.39-7.43 (m, 2H), 7.21-7.26(m, 1H), 1.70 (s, 6H).

Step 2:

To a mixture of 2-(3-bromophenyl)-2-methylpropanenitrile (37 g, 165mmol) in THE (500 mL) was added borane dimethylsulfide complex (49.5 mL,495 mmol) at 0° C. The resulting mixture was stirred at 30° C. for 12 h.The reaction was quenched with MeOH (200 mL) and HCl (12 M in water, 50mL) at 0° C. Then the mixture was stirred at 80° C. for 6 h. The mixturewas concentrated under reduced pressure to give to give2-(3-bromophenyl)-2-methylpropan-1-amine as colourless oil which wasused in the next step directly. MS: 228/230 (M+1/M+3).

Step 3:

To a mixture of 2-(3-bromophenyl)-2-methylpropan-1-amine (35 g, 153mmol) and TEA (64.2 mL, 460 mmol) in DCM (300 mL) was added2,5-dioxopyrrolidin-1-yl methyl carbonate (31.9 g, 184 mmol) at 28° C.The mixture was stirred at 28° C. for 1 h. The mixture was concentratedunder reduced pressure. The residue was purified by flash silica gelchromatography (eluting with ethyl acetate/pet. ether gradient) to givemethyl (2-(3-bromophenyl)-2-methylpropyl)carbamate as light brown oil.MS: 286/288 (M+1/M+3). ¹H NMR (400 MHz, CDCl₃) δ 7.46 (s, 1H), 7.36 (d,J=7.43 Hz, 1H), 7.28 (br s, 1H), 7.17-7.24 (m, 1H), 4.38 (s, 1H), 3.62(s, 3H), 3.36 (br d, J=6.6 Hz, 2H), 1.31 (s, 6H).

Step 4:

A mixture of methyl (2-(3-bromophenyl)-2-methylpropyl)carbamate (34 g,119 mmol) and trifluoromethanesulfonic acid (526 mL, 5941 mmol) wasstirred at 75° C. for 12 h. The mixture was slowly added into ice water(4 L) and extracted with DCM (3 L×3). The combined organic layers werewashed with brine (2 L), dried over anhydrous Na₂SO₄, filtered, andconcentrated. The residue was purified by Prep-HPLC (ACN/water with 0.1%TFA modifier) to give6-bromo-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one as a white solid.MS: 254/256 (M+1/M+3). ¹H NMR (400 MHz, CD₃OD) δ 7.86 (d, J=8.36 Hz,1H), 7.61 (d, J=1.76 Hz, 1H), 7.54 (dd, J=1.98, 8.14 Hz, 1H), 3.31 (s,2H), 1.34-1.37 (m, 6H).

Step 5:

6-bromo-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one (1 g, 3.94 mmol)was added to a well stirred solution of sodium hydride (0.236 g, 5.90mmol) in DMF (80 mL). The reaction mixture was allowed to stir at 20° C.for 0.5 h. Then (3S)-tert-butyl3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (1.192 g, 4.33mmol) was added to the mixture. The reaction mixture was stirred at 40°C. for 12 h. The mixture was slowly added into water (1.5 L) andextracted with EtOAc (800 mL×3). The combined organic layers were washedwith brine (1 L), dried over anhydrous Na₂SO₄, filtered, andconcentrated. The residue was purified by Prep-HPLC (ACN/water with 0.1%TFA modifier) to give(10aS)-1-((6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneas yellow oil. MS: 455/457 (M+1/M+3). ¹H NMR (400 MHz, CDCl₃) δ7.87-7.96 (m, 1H), 7.44-7.49 (m, 1H), 7.20-7.24 (m, 2H), 7.12-7.18 (m,3H), 4.73-4.84 (m, 2H), 4.30-4.45 (m, 2H), 4.06-4.14 (m, 1H), 3.78-3.85(m, 1H), 3.35-3.73 (m, 2H), 2.91-3.13 (m, 2H), 1.31-1.40 (m, 6H).

Step 6:

To a mixture of(10aS)-1-((6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-one(2.5 g, 5.49 mmol) in MeOH (20 mL) and water (20 mL) was added sodiumhydroxide (0.439 g, 10.98 mmol) at 20° C., then the solution was stirredat 75° C. for 36 h. The mixture was concentrated under reduced pressureto give6-bromo-2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(1.8 g) as light brown solid which was used in next step withoutpurification. MS: 429/431 (M+1/M+3).

Step 7:

To a solution of6-bromo-2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(1.8 g, 4.19 mmol) and TEA (1.169 mL, 8.38 mmol) in DCM (30 mL) wasadded (Boc)₂O (1.460 mL, 6.29 mmol) at 20° C. The mixture was stirred at20° C. for 1 h. The mixture was concentrated under reduced pressure. Theresidue was purified by flash silica gel chromatography (eluting withethyl acetate/pet. ether gradient) to give (3S)-tert-butyl3-(2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas a light yellow solid. MS: 529/531 (M+1/M+3).

Step 8:

The mixture of two stereoisomers was purified by chiral SFC (AD column,45%/55% ethanol/CO₂) to afford (S)-tert-butyl3-((R)-2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(first eluting): ¹H NMR (400 MHz, CD₃OD) δ 7.74 (br d, J=7.89 Hz, 1H),7.50 (d, J=1.32 Hz, 1H), 7.44 (dd, J=1.75, 8.33 Hz, 1H), 7.14 (s, 4H),4.22-4.63 (m, 2H), 3.97-4.19 (m, 1H), 3.77-3.95 (m, 1H), 3.53-3.68 (m,1H), 3.32-3.41 (m, 2H), 3.12-3.25 (m, 1H), 2.99-3.13 (m, 1H), 2.88-2.94(m, 1H), 1.44-1.58 (m, 9H), 1.18-1.32 (m, 6H) and (S)-tert-butyl3-((S)-2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(second eluting): ¹H NMR (400 MHz, CD₃OD) δ 7.81 (br s, 1H), 7.44-7.62(m, 2H), 7.05-7.23 (m, 4H), 4.78-4.82 (m, 1H), 4.25-4.45 (m, 2H),3.70-3.91 (m, 2H), 3.36-3.63 (m, 3H), 2.97-3.09 (m, 2H), 1.38-1.54 (m,9H), 1.24-1.33 (m, 6H).

Intermediate 10: (S)-tert-butyl3-((R)-2-(6′-chloro-1′-oxo-1′H-spiro[cyclobutane-1,4′-isoquinolin]-2′(3′H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate

Step 1:

Into a 1000-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of N₂, was placed 4-chloro-2-methylbenzoic acid (60 g,352 mmol), MeOH (600 ml) and sulfuric acid (37.5 ml, 703 mmol). Themixture was heated to 75° C. for 6 hours. The resulting solution wasquenched by adding 2 L of water and then extracted with 2×500 mL ofEtOAc. The organic layers were combined. The resulting mixture waswashed with 2×300 mL of brine, dried over anhydrous sodium sulfate, andconcentrated under vacuum to give methyl 4-chloro-2-methylbenzoate. ¹HNMR (300 MHz, CDCl₃) δ 7.86 (d, J=8.3 Hz, 1H), 7.29-7.14 (m, 2H), 3.89(s, 3H), 2.59 (s, 3H).

Step 2:

Into a 1000-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of N₂, was placed CCl₄ (600 ml), methyl4-chloro-2-methylbenzoate (60 g, 325 mmol), NBS (87 g, 487 mmol) and BPO(3.15 g, 13.00 mmol). The mixture was heated to 80° C. for 20 hours. Theresulting solution was cooled to room temperature. The mixture wasconcentrated under vacuum and then purified by HPLC to give methyl2-(bromomethyl)-4-chlorobenzoate. ¹H NMR (300 MHz, CDCl₃) δ 7.92 (d,J=8.4 Hz, 1H), 7.47 (d, J=2.2 Hz, 1H), 7.35 (dd, J=8.5, 2.2 Hz, 1H),4.91 (s, 2H), 3.94 (s, 3H).

Step 3:

Into a 1000-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of N₂, was placed methyl2-(bromomethyl)-4-chlorobenzoate (41 g, 156 mmol), DMSO (600 ml, 8455mmol) and water (60 ml). Then sodium cyanide (8.39 g, 171 mmol) wasadded in portions over 30 minutes at room temperature. The mixture wasstirred for 2 hours. The resulting solution was quenched by adding 1000mL aqueous iron(II) sulfate, and extracted with 3×500 mL EtOAc. Thecombined organic layers were washed with 2×300 mL of brine, dried overanhydrous sodium sulfate, and concentrated under vacuum. The resultingresidue was purified by HP-HPLC to give methyl4-chloro-2-(cyanomethyl)benzoate. ¹H NMR (300 MHz, CDCl₃) δ 8.02 (d,J=8.4 Hz, 1H), 7.59 (d, J=2.1 Hz, 1H), 7.48-7.36 (m, 1H), 4.26-4.14 (m,2H), 3.92 (s, 3H).

Step 4:

Into a 500-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of N₂, was placed methyl4-chloro-2-(cyanomethyl)benzoate (11 g, 52.5 mmol), acetonitrile (220ml), 1,3-diiodopropane (31 g, 105 mmol) and cesium carbonate (42.8 g,131 mmol). The mixture was heated to 75° C. for 20 hours. The resultingsolution was cooled to room temperature. The mixture was then filteredand the filtrate was concentrated under vacuum. The resulting residuewas purified by HPLC to give methyl4-chloro-2-(1-cyanocyclobutyl)benzoate. ¹H NMR (300 MHz, CDCl₃) δ 7.86(d, J=8.3 Hz, 1H), 7.38 (dd, J=8.3, 2.1 Hz, 1H), 7.33 (d, J=2.1 Hz, 1H),3.93 (s, 3H), 3.01-2.87 (m, 2H), 2.63-2.38 (m, 3H), 2.01-1.87 (m, 1H).

Step 5:

Into a 250-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of N₂, was placed methyl4-chloro-2-(1-cyanocyclobutyl)benzoate (7 g, 28 mmol), and THE (140 ml).After cooling to 0° C., Schwartz's reagent (14.4 g, 56 mmol) was added.The mixture was stirred for 2 hours at 0° C. The resulting solution wasquenched by adding 1500 mL aqueous ammonium chloride, and extracted with3×100 mL EtOAc. The organic layers were combined, washed with 2×100 mLof brine, dried over anhydrous sodium sulfate, and concentrated undervacuum. The resulting residue was purified by HPLC to give methyl4-chloro-2-(1-formylcyclobutyl)benzoate. MS: 253 (M+1). ¹H NMR (400 MHz,CDCl₃) δ 9.87 (s, 1H), 7.82 (d, J=8.2 Hz, 1H), 7.35 (dd, J=8.2, 2.1 Hz,1H), 7.32 (d, J=2.0 Hz, 1H), 3.84 (s, 3H), 2.76 (dddd, J=11.2, 6.7, 3.8,1.8 Hz, 2H), 2.45-2.34 (m, 2H), 2.15 (dq, J=11.6, 9.0 Hz, 1H), 1.99-1.85(m, 1H).

Step 6:

Into a 100-mL 3-necked round-bottom flask purged and maintained with aninert atmosphere of nitrogen, was placed a solution of methyl4-chloro-2-(1-formylcyclobutyl)benzoate (1.7 g, 6.73 mmol), DCM (50 ml),and (S)-tert-butyl3-((R)-2-amino-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(2.95 g, 10.1 mmol) at room temperature. Tri-isopropoxytitanium chloride(4.03 mmol, 1M in DCM), acetic acid (0.039 ml, 0.67 mmol), and sodiumtriacetoxyborohydride (7.13 g, 33.7 mmol) were then added sequentially.The resulting solution was stirred for 20 h at 25° C. The reaction wasthen quenched by the addition of 50 mL water and extracted with 2×100 mLof DCM. The organic layers were combined, washed with 2×50 mL of brine,dried over anhydrous sodium sulfate, and concentrated under vacuum. Theresulting residue was purified by silica gel chromatography to give(S)-tert-butyl3-((R)-2-(6′-chloro-1′-oxo-1′H-spiro[cyclobutane-1,4′-isoquinolin]-2′(3′H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas a white solid. MS: 497 (M+1). ¹H NMR (300 MHz, CD₃OD) δ 7.83 (d,J=8.3 Hz, 1H), 7.56 (d, J=2.0 Hz, 1H), 7.31 (dd, J=8.4, 2.1 Hz, 1H),7.16 (s, 4H), 4.94-4.85 (m, 1H), 4.63-4.39 (m, 1H), 4.33 (s, 1H),4.17-4.06 (m, 1H), 3.87 (m, 2H), 3.65 (m, 1H), 3.25-3.04 (m, 2H), 2.93(m, 1H), 2.36-2.17 (m, 3H), 2.08 (m, 3H), 1.52 (s, 9H).

Examples 1-4 Example 1:6-bromo-4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-3,4-dihydroisoquinolin-1(2H)-oneIsomer 1 Example 2:6-bromo-4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-3,4-dihydroisoquinolin-1(2H)-oneIsomer 2 Example 3:6-bromo-4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-3,4-dihydroisoquinolin-1(2H)-oneIsomer 3 Example 4:6-bromo-4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-3,4-dihydroisoquinolin-1(2H)-oneIsomer 4

Each of the following structures corresponds to one of theabove-referenced isomers:

A mixture of (3S)-tert-butyl3-(2-(6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 1 (50 mg, 0.094 mmol) and HCl (3 mL of 4N solution in EtOAc) wasstirred at 18° C. for 30 minutes. The mixture was concentrated underreduced pressure. The residue was purified by HPLC (ACN/water with 0.1%ammonium bicarbonate modifier) to give6-bromo-4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-3,4-dihydroisoquinolin-1(2H)-oneIsomer 1 (Example 1) as a white solid. MS: 429/431 (M+1/M+3). H NMR (400MHz, CD₃OD) δ 7.83 (d, J=8.33 Hz, 1H), 7.43-7.58 (m, 2H), 6.97-7.16 (m,4H), 3.89-4.08 (m, 5H), 3.62-3.73 (m, 1H), 3.56-3.59 (m, 1H), 2.76-2.98(m, 4H), 1.57-1.79 (m, 2H), 0.99 (t, J=7.45 Hz, 3H).

The other three isomers were prepared using the methodology herein andthe general procedure above.

Example 2: MS: 429/431 (M+1/M+3). ¹H NMR (400 MHz, CD₃OD) δ 7.83 (d,J=8.33 Hz, 1H), 7.44-7.56 (m, 2H), 6.97-7.17 (m, 4H), 3.81-4.10 (m, 5H),3.59-3.75 (m, 2H), 2.74-2.95 (m, 4H), 1.57-1.83 (m, 2H), 1.00 (t, J=7.45Hz, 3H).

Example 3: MS: 429/431 (M+1/M+3). ¹H NMR (400 MHz, CD₃OD) δ 7.83 (d,J=8.33 Hz, 1H), 7.45-7.60 (m, 2H), 7.00-7.17 (m, 4H), 3.96-4.09 (m, 2H),3.79-3.93 (m, 3H), 3.55-3.75 (m, 2H), 2.71-2.99 (m, 4H), 1.57-1.81 (m,2H), 1.00 (t, J=7.45 Hz, 3H).

Example 4: MS: 429/431 (M+1/M+3). ¹H NMR (400 MHz, CD₃OD) δ 7.83 (d,J=8.33 Hz, 1H), 7.44-7.56 (m, 2H), 6.99-7.18 (m, 4H), 3.95-4.05 (m, 4H),3.81-3.89 (m, 1H), 3.51-3.65 (m, 1H), 3.49 (m, 1H), 2.73-2.95 (m, 4H),1.62-1.78 (m, 2H), 0.90-1.07 (m, 3H).

Examples 5-8 Example 5:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(1-methyl-1H-1,2,4-triazol-5-yl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 1 Example 6:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(1-methyl-1H-1,2,4-triazol-5-yl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 2 Example 7:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(1-methyl-1H-1,2,4-triazol-5-yl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 3 Example 8:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(1-methyl-1H-1,2,4-triazol-5-yl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 4

Each of the following structures corresponds to one of theabove-referenced isomers:

Step 1:

A mixture of(10aS)-1-((6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-one(300 mg, 0.659 mmol), 1-methyl-1H-1,2,4-triazole (109 mg, 1.318 mmol),2,2-dimethylbutanoic acid (22.96 mg, 0.198 mmol),di(adamantan-1-yl)(butyl)phosphine (94 mg, 0.264 mmol), potassiumcarbonate (364 mg, 2.64 mmol) and palladium(II) acetate (29.6 mg, 0.132mmol) in toluene (6 ml) was degassed and backfilled with N₂ (threetimes). The mixture was heated to 120° C. for 12 h. The mixture wascooled and concentrated under reduced pressure. The residue wasdissolved in water (10 mL) and EtOAc (10 mL). The organic layer wasseparated and the aqueous was re-extracted with EtOAc (10 mL×3). Thecombined organic layers were washed with brine (20 mL), dried overanhydrous Na₂SO₄, filtered, and concentrated. The residue was purifiedby flash silica gel chromatography (eluting with ethyl acetate/pet.ether gradient) to give(10aS)-1-((4-ethyl-6-(1-methyl-1H-1,2,4-triazol-5-yl)-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneas a white solid. MS: 458 (M+1). H NMR (400 MHz, CDCl₃) δ 8.11-8.26 (m,1H), 7.90-7.98 (m, 1H), 7.61-7.70 (m, 1H), 7.56-7.61 (m, 1H), 7.07-7.24(m, 4H), 4.73-4.95 (m, 1H), 4.55-4.71 (m, 1H), 4.29-4.46 (m, 1H),4.06-4.22 (m, 1H), 3.99-4.05 (m, 3H), 3.90-3.97 (m, 1H), 3.60-3.87 (m,2H), 3.00-3.27 (m, 1H), 2.77-2.97 (m, 3H), 1.54-1.65 (m, 2H), 0.96-1.07(m, 3H).

Step 2:

The above mixture of four stereoisomers was purified by chiral SFC (ADcolumn, 40%/60% methanol/CO₂) to afford four isomers:(10aS)-1-((4-ethyl-6-(1-methyl-1H-1,2,4-triazol-5-yl)-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneIsomer 1 (first eluting) and(10aS)-1-((4-ethyl-6-(1-methyl-1H-1,2,4-triazol-5-yl)-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneIsomer 2 (second eluting) and(10aS)-1-((4-ethyl-6-(1-methyl-1H-1,2,4-triazol-5-yl)-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneIsomer 3 (third eluting) and(10aS)-1-((4-ethyl-6-(1-methyl-1H-1,2,4-triazol-5-yl)-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneIsomer 4 (fourth eluting).

Step 3:

To a mixture of(10aS)-1-((4-ethyl-6-(1-methyl-1H-1,2,4-triazol-5-yl)-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneIsomer 1 (15 mg, 0.033 mmol) in MeOH (1 mL) and water (1 mL) was addedsodium hydroxide (2.62 mg, 0.066 mmol) at 20° C. Then the mixture wasstirred at 60° C. for 60 h. The mixture was concentrated under reducedpressure. The residue was purified by IPLC (ACN/water with 0.1% ammoniumbicarbonate modifier) to give4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(1-methyl-1H-1,2,4-triazol-5-yl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 1 (Example 5) as a white solid. MS: 432 (M+1). ¹H NMR (400 MHz,CDCl₃) δ 8.19 (d, J=7.89 Hz, 1H), 7.96 (s, 1H), 7.52-7.63 (m, 2H), 7.15(d, J=2.19 Hz, 3H), 7.04 (br d, J=4.82 Hz, 1H), 3.99-4.10 (m, 5H),3.88-3.99 (m, 2H), 3.64-3.83 (m, 3H), 2.76-3.02 (m, 4H), 1.73-1.82 (m,2H), 1.02 (t, J=7.24 Hz, 3H).

The other three isomers were prepared using the methodology herein andthe general procedure above.

Example 6: MS: 432 (M+1). ¹H NMR (400 MHz, CD₃OD) δ 8.13 (d, J=8.33 Hz,1H), 8.02 (s, 1H), 7.62-7.78 (m, 2H), 6.96-7.19 (m, 4H), 3.94-4.11 (m,8H), 3.54-3.79 (m, 2H), 2.82-3.03 (m, 4H), 1.73-1.80 (m, 2H), 1.04 (t,J=7.45 Hz, 3H).

Example 7: MS: 432 (M+1). ¹H NMR (400 MHz, CD₃OD) δ 8.13 (d, J=7.89 Hz,1H), 8.02 (s, 1H), 7.73 (dd, J=1.75, 7.89 Hz, 1H), 7.67 (s, 1H),7.03-7.14 (m, 4H), 3.87-4.09 (m, 8H), 3.68-3.78 (m, 2H), 2.87-2.99 (m,4H), 1.71-1.85 (m, 2H), 1.04 (t, J=7.45 Hz, 3H).

Example 8: MS: 432 (M+1). H NMR (400 MHz, CD₃OD) δ 8.13 (d, J=7.89 Hz,1H), 8.02 (s, 1H), 7.73 (dd, J=1.53, 8.11 Hz, 1H), 7.68 (s, 1H),6.99-7.20 (m, 4H), 3.98-4.15 (m, 7H), 3.82-3.92 (m, 1H), 3.65-3.74 (m,1H), 3.49-3.55 (m, 1H), 2.72-3.05 (m, 4H), 1.73-1.80 (m, 2H), 1.04 (t,J=7.45 Hz, 3H).

Example 96-(1-cyclopropyl-1H-1,2,4-triazol-5-yl)-2-{(2R)-2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one

Step 1:

To an oven-dried, nitrogen-cooled via was added S-ter-butyl3-((R)-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(100 mg, 0.189 mmol), bis(pinacolato)diboron (96 mg, 0.38 mmol),potassium acetate (37 mg, 0.38 mmol), and1,1′-bis(di-tert-butylphosphino)ferrocene palladium dichloride (12 mg,0.019 mmol). DMA (1 mL) was added, and the mixture was degassed for 10min by streaming nitrogen through the mixture. The reaction mixture wasstirred for 18 h at 90° C. The reaction was cooled to room temperature,diluted with water, filtered over Celite, and extracted with EtOAc (3×).The combined organic layers were washed with brine, dried over anhydroussodium sulfate, filtered, and concentrated under reduced pressure. Theresidue was purified by column chromatography on silica (eluting withethyl acetate/pet. ether gradient) to afford a ˜2:1 mixture of(S)-tert-butyl3-((R)-2-(4,4-dimethyl-1-oxo-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateand(2-((R)-2-((S)-2-(tert-butoxycarbonyl)-1,2,3,4-tetrahydroisoquinolin-3-yl)-2-hydroxyethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinolin-6-yl)boronicacid as a solid. MS: 577 (M+1) and 495 (M+1), respectively.

Step 2:

To an oven-dried, nitrogen-cooled vial was added5-bromo-1-cyclopropyl-1H-1,2,4-triazole (19 mg, 0.10 mmol), a ˜2:1mixture of (S)-tert-butyl3-((R)-2-(4,4-dimethyl-1-oxo-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateand(2-((R)-2-((S)-2-(tert-butoxycarbonyl)-1,2,3,4-tetrahydroisoquinolin-3-yl)-2-hydroxyethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinolin-6-yl)boronicacid (54 mg, 0.094 mmol),(2-dicyclohexylphosphino-2′,4′,6′-triisopropyl-1,1′-biphenyl)[2-(2′-amino-1,1′-biphenyl)]palladium(II)methanesulfonate (7.9 mg, 9.4 μmol), THE (1 mL), and potassiumphosphate, tribasic (1.0 M in water, 470 μl, 0.470 mmol). The mixturewas purged with nitrogen and heated at 80° C. for 18 h. The mixture wascooled to room temperature, diluted with water (10 mL), and extractedwith EtOAc (2×20 mL). The combined organic layers were washed with brine(20 mL), dried over anhydrous sodium sulfate, filtered, and concentratedunder reduced pressure. The residue was dissolved in DMSO (2 mL),filtered, and purified by mass triggered reverse phase HPLC (ACN/waterwith 0.1% TFA modifier) to afford (S)-tert-butyl3-((R)-2-(6-(1-cyclopropyl-1H-1,2,4-triazol-5-yl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate,TFA as a solid. MS: 558 (M+1).

Step 3:

To a solution of (S)-tert-butyl3-((R)-2-(6-(1-cyclopropyl-1H-1,2,4-triazol-5-yl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate,TFA (35 mg, 0.052 mmol) in DCM (1.5 mL) was added HCl (4.0 M in dioxane,64 μl, 0.26 mmol). The mixture was stirred at room temperature for 1 h,and then another 5 eq of HCl was added and the mixture was stirred for 2h at room temperature. The mixture was concentrated under reducedpressure to afford6-(1-cyclopropyl-1H-1,2,4-triazol-5-yl)-2-{(2R)-2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one,HCl as a solid. MS 458 (M+1). ¹H NMR (400 MHz, CD₃OD) δ: 8.61 (s, 1H),8.25 (d, J=7.4 Hz, 1H), 8.02 (s, 1H), 7.95 (d, J=7.7 Hz, 1H), 7.36-7.26(m, 3H), 7.26-7.22 (m, 1H), 4.55-4.34 (m, 4H), 4.09-3.98 (m, 2H),3.79-3.60 (m, 5H), 1.53-1.43 (m, 6H), 1.26-1.16 (m, 4H).

The compounds in the following table were prepared using the methodologyherein and the general procedure described in Example 9.

Ex. No. Structure Chemical Name [M + H]+ 10

  Enantiopure (using Isomer 2 of starting material)6-(1-cyclopropyl-1H-1,2,4- triazol-5-yl)-4-ethyl-2-{(2R)-2-hydroxy-2-[(3S)- 1,2,3,4- tetrahydroisoquinolin-3-yl]ethyl}-3,4- dihydroisoquinolin-1(2H)- one 458 11

6′-(1-cyclopropyl-1H-1,2,4- triazol-5-yl)-2′-{(2R)-2-hydroxy-2-[(3S)-1,2,3,4- tetrahydroisoquinolin-3-yl]ethyl}-2′,3′-dihydro-1′H- spiro[cyclobutane-1,4′- isoquinolin]-1′-one470

Example 124-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-one

Step 1:

To a solution of 6-bromo-4-ethyl-3,4-dihydroisoquinolin-1(2H)-one (400mg, 1.574 mmol) and potassium acetate (463 mg, 4.72 mmol) in ethanol (20mL) was added PdCl₂(dppf) (230 mg, 0.315 mmol) under N₂ atmosphere. Themixture was degassed and backfilled with CO (three times). The resultingmixture was stirred under CO (50 psi) at 75° C. for 5 h. The catalystwas filtered off and filtrate was concentrated under reduced pressure.The residue was purified by flash silica gel chromatography (elutingwith ethyl acetate/pet. ether gradient) to give ethyl4-ethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylate as a brownsolid. MS: 248 (M+1). H NMR (400 MHz, CDCl₃) δ 8.11 (d, J=8.02 Hz, 1H),7.99 (dd, J=1.57, 8.02 Hz, 1H), 7.88 (d, J=1.56 Hz, 1H), 6.29 (br s,1H), 4.33-4.44 (m, 2H), 3.70-3.72 (m, 1H), 3.39-3.46 (m, 1H), 2.77-2.85(m, 1H), 1.69-1.80 (m, 2H), 1.40 (t, J=7.14 Hz, 3H), 0.97 (t, J=7.43 Hz,3H).

Step 2:

To a solution of ethyl4-ethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylate (300 mg, 1.21mmol) in EtOH (5 mL) and water (2 mL) was added LiOH (29 mg, 1.21 mmol)at 30° C. The mixture was stirred at 30° C. for 1 h. The mixture wasconcentrated under reduced pressure. The residue was dissolved in water(10 mL) and acidified to pH 6 with HCl (2 M in water). The aqueous wasextracted with EtOAc (10 mL×3). The combined organic layers were washedwith brine (10 mL), dried over anhydrous Na₂SO₄, filtered, andconcentrated to give4-ethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylic acid as awhite solid which was used in the next step directly. MS: 220 (M+1).

Step 3:

To a solution of4-ethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylic acid (240 mg,1.095 mmol) in DMF (4 mL) was added HATU (624 mg, 1.642 mmol) and DIEA(0.574 mL, 3.28 mmol) at 25° C. The mixture was stirred for 5 minutes at25° C. under N₂. 3-oxa-8-azabicyclo[3.2.1]octane hydrochloride (197 mg,1.314 mmol) was added. The mixture was stirred at 25° C. for 1 h. Themixture was then dissolved in water (20 mL) and EtOAc (25 mL). Theorganic layer was separated and the aqueous was re-extracted with EtOAc(20 mL×3). The combined organic layers were washed with brine (30 mL),dried over anhydrous Na₂SO₄, filtered and concentrated. The residue waspurified by column chromatography on silica gel (eluting with ethylacetate/pet. ether gradient) to give6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-3,4-dihydroisoquinolin-1(2H)-oneas a brown solid. MS: 315 (M+1).

Step 4:

To a solution of NaH (22.90 mg, 0.573 mmol) in DMF (2 mL) was added6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-3,4-dihydroisoquinolin-1(2H)-one(120 mg, 0.382 mmol) at 20° C. The reaction mixture was stirred at 20°C. for 0.5 h. Then (3S)-tert-butyl3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (95 mg, 0.344mmol) was added to the mixture. The reaction mixture was stirred at 80°C. for 12 h. The residue was purified by HPLC (ACN/water with 0.1%NH₄HCO₃ modifier) to give4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-oneas a white solid. MS: 490 (M+1). ¹H NMR (400 MHz, CD₃OD) δ 8.02 (d,J=7.82 Hz, 1H), 7.38-7.49 (m, 2H), 7.01-7.20 (m, 4H), 4.50-4.78 (m, 1H),3.78-4.07 (m, 6H), 3.48-3.74 (m, 5H), 2.74-2.97 (m, 4H), 2.30-2.36 (m,1H), 1.92-2.12 (m, 4H), 1.61-1.82 (m, 2H), 0.99 (br s, 3H).

Example 13-16 Example 13:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 1 Example 14:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 2 Example 15:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 3, and Example 16:4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 4

Each of the following structures corresponds to one of theabove-referenced isomers:

Step 1:

To a solution of(10aS)-1-((6-bromo-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-one(1.1 g, 2.416 mmol) and potassium acetate (0.711 g, 7.25 mmol) inethanol (20 mL) was added PdCl₂(dppf) (0.354 g, 0.483 mmol) under N₂atmosphere. The mixture was degassed and backfilled with CO (threetimes). The resulting mixture was stirred under CO (50 psi) at 70° C.for 12 h. The mixture was concentrated under reduced pressure. Theresidue was purified by flash silica gel chromatography (eluting withethyl acetate/pet. ether gradient) to give ethyl4-ethyl-1-oxo-2-(((10aS)-3-oxo-3,5,10,10a-tetrahydro-1H-oxazolo[3,4-b]isoquinolin-1-yl)methyl)-1,2,3,4-tetrahydroisoquinoline-6-carboxylateas a brown solid. MS: 449 (M+1).

Step 2:

To a mixture of ethyl4-ethyl-1-oxo-2-(((10aS)-3-oxo-3,5,10,10a-tetrahydro-1H-oxazolo[3,4-b]isoquinolin-1-yl)methyl)-1,2,3,4-tetrahydroisoquinoline-6-carboxylate(800 mg, 1.784 mmol) in ethanol (5 mL) and water (2 mL) was addedlithium hydroxide hydrate (85 mg, 3.57 mmol) at 20° C. Then the mixturewas stirred at 20° C. for 6 h. The mixture was concentrated underreduced pressure. Water (20 mL) was added and the mixture was acidifiedto pH 5 with an aqueous citric acid solution. The mixture was extractedwith EtOAc (50 mL×3). The combined organic layers were washed with brine(60 mL), dried over anhydrous Na₂SO₄, filtered, and concentrated invacuum to give4-ethyl-1-oxo-2-(((10aS)-3-oxo-3,5,10,10a-tetrahydro-1H-oxazolo[3,4-b]isoquinolin-1-yl)methyl)-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid as a light yellow solid which would be used without furtherpurification. MS: 421 (M+1).

Step 3:

To a mixture of4-ethyl-1-oxo-2-(((10aS)-3-oxo-3,5,10,10a-tetrahydro-1H-oxazolo[3,4-b]isoquinolin-1-yl)methyl)-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid (720 mg, 1.712 mmol), HATU (846 mg, 2.226 mmol) and DIEA (0.897 mL,5.14 mmol) in DMF (10 mL) was added 3-oxa-8-azabicyclo[3.2.1]octanehydrochloride (282 mg, 1.884 mmol) at 20° C. Then the mixture wasstirred at 20° C. for 0.5 h. To the mixture was added water (100 mL) andEtOAc (50 mL). The organic layer was separated and the aqueous wasre-extracted with EtOAc (50 mL×3). The combined organic layers werewashed with brine (50 mL), dried over anhydrous Na₂SO₄, filtered, andconcentrated. The residue was purified by Prep-TLC (silica gel, ethylacetate:pet. ether=3:1, v/v) to give(10aS)-1-((6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneas brown oil. MS: 516 (M+1).

Step 4:

To a mixture of(10aS)-1-((6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-one(750 mg, 1.455 mmol) in MeOH (10 mL) and water (10 mL) was added NaOH(116 mg, 2.91 mmol) at 20° C. Then the mixture was stirred at 50° C. for48 h. The mixture was concentrated under reduced pressure. The residuewas dissolved in water (40 mL) and extracted with EtOAc (40 mL×2), thecombined organic layers were dried over Na₂SO₄, filtered, andconcentrated to give4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-oneas a white solid which was used in the next step directly withoutfurther purification. MS: 490 (M+1).

Step 5:

To a solution of4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-one(550 mg, 1.123 mmol) and TEA (0.470 mL, 3.37 mmol) in DCM (15 mL) wasadded (Boc)₂O (0.522 mL, 2.247 mmol). The mixture was stirred at 20° C.for 0.5 h. The mixture was concentrated under reduced pressure. Theresidue was purified by flash silica gel chromatography (eluting withethyl acetate/pet. ether gradient) to give (3S)-tert-butyl3-(2-(6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas a light yellow solid. MS: 590 (M+1).

Step 6:

The above mixture was purified by chiral SFC (AD column, 25%/75%methanol/CO₂) to afford four stereoisomers: (3S)-tert-butyl3-(2-(6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 1 (first eluting) and (3S)-tert-butyl3-(2-(6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 2 (second eluting) and (3S)-tert-butyl3-(2-(6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 3 (third eluting) and (3S)-tert-butyl3-(2-(6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 4 (fourth eluting).

Step 7:

A mixture of (3S)-tert-butyl3-(2-(6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4-ethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateIsomer 1 (40 mg, 0.068 mmol) and HCl (3 mL of a 4M solution in EtOAc)was stirred at 18° C. for 30 minutes. The mixture was concentrated underreduced pressure. The residue was purified by HPLC (ACN/water with 0.1%NH₄HCO₃ modifier) to give4-ethyl-2-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-oneIsomer 1 (Example 13) as a white solid. MS: 490 (M+1). ¹H NMR (400 MHz,CD₃OD) δ 7.97-8.08 (m, 1H), 7.36-7.53 (m, 2H), 7.00-7.17 (m, 4H), 4.65(br s, 1H), 3.90-4.09 (m, 6H), 3.78-3.86 (m, 1H), 3.65-3.76 (m, 3H),3.52-3.64 (m, 2H), 2.83-3.02 (m, 4H), 1.90-2.11 (m, 4H), 1.66-1.80 (m,2H), 0.92-1.07 (m, 3H).

The other three isomers were prepared using the methodology herein andthe general procedure above.

Example 14: MS: 490 (M+1). H NMR (400 MHz, CD₃OD) δ 8.04 (d, J=7.89 Hz,1H), 7.48 (dd, J=1.32, 7.89 Hz, 1H), 7.42 (s, 1H), 7.00-7.15 (m, 4H),4.58-4.73 (m, 1H), 3.77-4.11 (m, 7H), 3.64-3.76 (m, 4H), 3.54-3.60 (m,1H), 2.80-2.99 (m, 4H), 1.90-2.12 (m, 4H), 1.63-1.83 (m, 2H), 1.01 (brt, J=5.92 Hz, 3H).

Example 15: MS: 490 (M+1). H NMR (400 MHz, CD₃OD) δ 8.04 (d, J=7.94 Hz,1H), 7.38-7.53 (m, 2H), 7.00-7.18 (m, 4H), 4.65 (br s, 1H), 3.98-4.10(m, 2H), 3.78-3.96 (m, 5H), 3.61-3.76 (m, 4H), 3.54-3.60 (m, 1H),2.77-3.01 (m, 4H), 1.92-2.12 (m, 4H), 1.61-1.84 (m, 2H), 1.01 (br t,J=7.06 Hz, 3H).

Example 16: MS: 490 (M+1). ¹H NMR (400 MHz, CD₃OD) δ 8.04 (d, J=7.89 Hz,1H), 7.35-7.54 (m, 2H), 7.01-7.17 (m, 4H), 4.65 (br s, 1H), 3.91-4.10(m, 5H), 3.77-3.90 (m, 2H), 3.63-3.76 (m, 3H), 3.45-3.61 (m, 2H),2.74-2.96 (m, 4H), 1.93-2.13 (m, 4H), 1.60-1.83 (m, 2H), 1.00 (br t,J=6.80 Hz, 3H).

Examples 17-18 Example 17:6-bromo-2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-oneand Example 18:6-bromo-2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one

A mixture of (S)-tert-butyl3-((R)-2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(40 mg, 0.076 mmol) and HCl/EtOAc (4 N) (3.0 mL) was stirred at 18° C.for 30 minutes. The mixture was concentrated under reduced pressure. Theresidue was purified by HPLC (ACN/water with 0.1% NH₄HCO₃ modifier) togive6-bromo-2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(Example 17) as a white solid. MS: 429/431 (M+1/M+3). ¹H NMR (400 MHz,CD₃OD) δ 7.85 (d, J=8.33 Hz, 1H), 7.48-7.60 (m, 2H), 7.02-7.15 (m, 4H),3.88-4.08 (m, 4H), 3.50-3.68 (m, 3H), 2.82-2.95 (m, 3H), 1.32-1.38 (m,6H).

Example 18 was prepared using the methodology herein and the generalprocedure described above, starting (S)-tert-butyl3-((S)-2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate.Example 18:6-bromo-2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one:MS: 429/431 (M+1/M+3). H NMR (400 MHz, CD₃OD) δ 7.85 (d, J=8.33 Hz, 1H),7.58 (d, J=1.75 Hz, 1H), 7.52 (dd, J=2.19, 8.33 Hz, 1H), 7.01-7.16 (m,4H), 3.79-4.09 (m, 4H), 3.47-3.67 (m, 3H), 2.74-2.95 (m, 3H), 1.36 (d,J=5.26 Hz, 6H).

Examples 19-20 Example 19:6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-oneand Example 20:6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one

Step 1:

To a solution of 6-bromo-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(2.55 g, 10.03 mmol) in ethanol (30 mL) was added potassium acetate(2.95 g, 30.1 m mol) and PdCl₂(dppf) (0.734 g, 1.003 mmol) under N₂atmosphere. The mixture was degassed and backfilled with CO (threetimes). The resulting mixture was stirred under CO (50 psi) at 75° C.for 12 h. The reaction mixture was concentrated under reduced pressureand the residue was purified by column chromatography on silica (elutingwith ethyl acetate/pet. ether gradient) to afford ethyl4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylate as acolorless oil. MS: 248 (M+1), ¹H NMR (400 MHz, CD₃OD) δ 8.07-8.02 (m,2H), 7.99 (d, J=1.5 Hz, 1H), 4.40 (q, J=7.1 Hz, 2H), 3.32 (br s, 2H),1.43-1.39 (m, 3H), 1.40-1.37 (m, 6H).

Step 2:

To a solution of ethyl4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylate (1 g,4.04 mmol) in MeOH (3 mL) and H₂O (2 mL) was added lithium hydroxidehydrate (0.204 g, 4.85 mmol) in 30° C. The reaction mixture was stirredat 30° C. for 4 h. The reaction was concentrated under reduced pressureto give 4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid which was used directly for the next step without purification. MS:220 (M+1).

Step 3:

To a solution4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylic acid (660mg, 3.01 mmol) in DCM (5 mL) was added2-(3H-[1,2,3]triazolo[4,5-b]pyridin-3-yl)-1,1,3,3-tetramethylisouroniumhexafluorophosphate(V)(1374 mg, 3.61 mmol), N-ethyl-N-isopropylpropan-2-amine (1.577 mL, 9.03mmol) and N-ethyl-N-isopropylpropan-2-amine (1.577 mL, 9.03 mmol) at 20°C. The resulting mixture stirred at 20° C. for 0.5 h. The reactionmixture was concentrated under reduced pressure and the residue waspurified by column chromatography on silica (eluting with ethylacetate/pet. ether gradient) to afford6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-oneas a yellow oil. MS: 315 (M+1). ¹H NMR (400 MHz, CDCl₃) δ 8.12 (d, J=7.7Hz, 1H), 7.50 (d, J=1.3 Hz, 1H), 7.41 (dd, J=1.5, 7.9 Hz, 1H), 3.72-3.92(m, 5H), 3.35 (s, 2H), 3.19 (dd, J=4.5, 7.4 Hz, 1H), 2.01-2.05 (m, 4H),1.39 (s, 6H).

Step 4:

To a mixture of sodium hydride (28.6 mg, 0.716 mmol) in DMF (2 mL) wasadded6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(150 mg, 0.477 mmol) at 20° C. The solution was stirred at 20° C. for0.5 h. To the resulting mixture was then added a solution of(3S)-tert-butyl3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (158 mg, 0.573mmol) in DMF (1 mL). The mixture was stirred at 80° C. for 15 h. Thereaction mixture was concentrated under reduced pressure and the residuewas purified by prep-HPLC (ACN/water with 0.1% ammonia hydroxidemodifier) to afford6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-oneas a light yellow oil. MS: 490 (M+1).

Step 5:

The above mixture of two stereoisomers was purified by chiral SFC (ICcolumn, 5%/40% ethanol/CO₂) to afford:6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-oneas a white solid (Example 19) (first eluting): MS: 490 (M+1). H NMR (400MHz, CD₃OD) δ 8.08 (d, J=7.9 Hz, 1H), 7.56 (d, J=1.2 Hz, 1H), 7.49 (dd,J=1.4, 7.9 Hz, 1H), 7.10-7.18 (m, 3H), 7.04-7.09 (m, 1H), 4.68 (br s,2H), 4.01-4.12 (m, 3H), 3.92-4.01 (m, 2H), 3.81-3.89 (m, 1H), 3.71-3.78(m, 2H), 3.65-3.71 (m, 1H), 3.54-3.65 (m, 2H), 2.88-3.00 (m, 3H), 2.04(br d, J=8.7 Hz, 4H), 1.41 (d, J=1.2 Hz, 6H) and6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(Example 20) (Second eluting): MS: 490 (M+1). ¹H NMR (400 MHz, CD₃OD)δ=8.06 (d, J=7.9 Hz, 1H), 7.55 (d, J=1.2 Hz, 1H), 7.48 (dd, J=1.5, 7.9Hz, 1H), 7.10-7.19 (m, 3H), 7.08 (br d, J=5.1 Hz, 1H), 4.11 (s, 1H),4.07 (d, J=6.4 Hz, 2H), 3.98-4.03 (m, 1H), 3.98-4.03 (m, 1H), 3.81-3.92(m, 2H), 3.68-3.76 (m, 2H), 3.63-3.67 (m, 2H), 3.53-3.62 (m, 2H),2.81-2.98 (m, 3H), 1.96-2.14 (m, 4H), 1.40 (d, J=4.6 Hz, 6H).

Examples 21-22 Example 21:2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3R,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-oneand Example22:2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3S,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one

Step 1:

To a solution of NaH (0.236 g, 5.90 mmol) in DMF (40 mL) was added6-bromo-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one (1.0 g, 3.94 mmol)at 20° C. The reaction mixture was allowed to stir at 20° C. for 0.5 h.Then (3S)-tert-butyl3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (1.192 g, 4.33mmol) was added to the mixture. The reaction mixture was stirred at 40°C. for 12 h. The reaction was poured into ice water (400 mL) andextracted with EtOAc (200 mL×2). The combined organic layers were driedover Na₂SO₄, filtered, and concentrated in vacuum. The residue waspurified by HPLC (ACN/water with 0.1% TFA modifier) to give(10aS)-1-((6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-oneas white solid. MS: 455/457 (M+1/M+3). ¹H NMR (400 MHz, CDCl₃) δ7.87-7.95 (m, 1H), 7.44-7.52 (m, 2H), 7.10-7.27 (m, 4H), 4.74-5.05 (m,2H), 4.64 (dt, J=2.9, 6.1 Hz, 1H), 4.32-4.34 (m, 1H), 4.07-4.16 (m, 1H),3.77-3.84 (m, 1H), 3.58-3.75 (m, 1H), 3.37-3.55 (m, 1H), 3.08-3.21 (m,1H), 2.86-3.00 (m, 1H), 1.30-1.45 (m, 6H).

Step 2:

A mixture of KOAc (582 mg, 5.93 mmol),(10aS)-1-((6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)methyl)-10,10a-dihydro-1H-oxazolo[3,4-b]isoquinolin-3(5H)-one(900 mg, 1.977 mmol) and Pd(dppf)Cl₂ (145 mg, 0.198 mmol) in EtOH (20ml) was degassed and backfilled with CO (three times). The resultingmixture was stirred under 50 psi of CO at 80° C. for 12 h. The mixturewas concentrated in vacuum. The residue was purified by flash silica gelchromatography (eluting with ethyl acetate/pet. ether gradient) to giveethyl4,4-dimethyl-1-oxo-2-(((10aS)-3-oxo-3,5,10,10a-tetrahydro-1H-oxazolo[3,4-b]isoquinolin-1-yl)methyl)-1,2,3,4-tetrahydroisoquinoline-6-carboxylateas an oil. MS: 449 (M+1). H NMR (400 MHz, CDCl₃) δ 8.09-8.19 (m, 1H),7.95-8.03 (m, 2H), 7.08-7.25 (m, 4H), 4.78 (s, 2H), 4.64 (dt, J=3.0, 6.0Hz, 1H), 4.29-4.46 (m, 3H), 4.06-4.12 (m, 1H), 3.78-3.89 (m, 1H), 3.63(s, 1H), 3.38-3.58 (m, 1H), 3.10-3.22 (m, 1H), 2.87-3.01 (m, 1H),1.31-1.50 (m, 9H).

Step 3:

A mixture of ethyl4,4-dimethyl-1-oxo-2-(((10aS)-3-oxo-3,5,10,10a-tetrahydro-1H-oxazolo[3,4-b]isoquinolin-1-yl)methyl)-1,2,3,4-tetrahydroisoquinoline-6-carboxylate(750 mg, 1.67 mmol) and sodium hydroxide (268 mg, 6.7 mmol) in MeOH (5ml) and water (5 ml) was stirred at 70° C. for 18 h. The mixture wasacidified to pH 5 with 1 N aqueous HCl and extracted with EtOAc. Theorganic layers were dried over Na₂SO₄, filtered and concentrated toafford2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid which was used without further purification. MS: 395 (M+1).

Step 4:

To a mixture of2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid (660 mg, 1.673 mmol) and sodium hydroxide (66.9 mg, 1.67 mmol) inMeOH (5 ml) and Water (5 ml) was added (Boc)₂O (0.971 ml, 4.18 mmol).The mixture was stirred for 30 min at 15° C. and then concentrated invacuo. The resulting residue was purified by HPLC (ACN/water with 0.1%TFA modifier) to give2-(2-((S)-2-(tert-butoxycarbonyl)-1,2,3,4-tetrahydroisoquinolin-3-yl)-2-hydroxyethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid as a brown solid. MS: 495 (M+1). ¹H NMR (400 MHz, CDCl₃) δ7.70-8.15 (m, 3H), 7.07-7.24 (m, 4H), 4.64-4.86 (m, 1H), 4.17-4.56 (m,3H), 3.73-4.02 (m, 2H), 3.32-3.61 (m, 3H), 2.95-3.20 (m, 2H), 1.45-1.54(m, 9H), 1.30-1.33 (m, 6H).

Step 5:

To a mixture of2-(2-((S)-2-(tert-butoxycarbonyl)-1,2,3,4-tetrahydroisoquinolin-3-yl)-2-hydroxyethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid (100 mg, 0.202 mmol), (1R,3r,5S)-8-azabicyclo[3.2.1]octan-3-olhydrochloride (39.7 mg, 0.243 mmol) and DIEA (0.106 ml, 0.607 mmol) inDMF (2 ml) was added 2,4,6-tripropyl-1,3,5,2,4,6-trioxatriphosphinane2,4,6-trioxide (257 mg, 0.404 mmol). The mixture was stirred for 1 h at15° C. Water (15 mL) was added to the mixture, and the mixture wasextracted with EtOAc (10 mL×3). The combined organic layers wereconcentrated in vacuum to give tert-butyl(3S)-3-(1-hydroxy-2-(6-((1R,3S,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas an oil, which was used in the next step without further purification.MS: 504 (M−Boc+H). ¹H NMR (400 MHz, CDCl₃) δ 8.03-8.10 (m, 1H),7.32-7.41 (m, 2H), 7.19 (dd, J=3.3, 5.4 Hz, 3H), 7.12 (br d, J=3.8 Hz,1H), 4.77-4.84 (m, 1H), 4.30-4.44 (m, 2H), 4.18-4.24 (m, 1H), 3.93-4.00(m, 1H), 3.74-3.88 (m, 2H), 3.47 (br s, 4H), 3.03-3.16 (m, 2H),2.20-2.37 (m, 3H), 1.99 (br s, 4H), 1.79 (br s, 1H), 1.45-1.52 (m, 9H),1.24-1.39 (m, 6H).

Step 6:

tert-butyl(3S)-3-(1-hydroxy-2-(6-((1R,3S,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(110 mg, 0.182 mmol) was separated by SFC to afford two isomers:tert-butyl(S)-3-((R)-1-hydroxy-2-(6-((1R,3R,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(first eluting) and tert-butyl(S)-3-((S)-1-hydroxy-2-(6-((1R,3S,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(second eluting).

Step 7:

A solution of tert-butyl(S)-3-((R)-1-hydroxy-2-(6-((1R,3R,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(30 mg, 0.050 mmol) in HCl (4 ml of 1 N in EtOAc) was stirred at 15° C.for 4 h. The mixture was concentrated in vacuo. The resulting residuewas purified by HPLC (ACN/water with 10 mM NH₄HCO₃ modifier) to give2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3R,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(Example. 21) as a white solid. MS: 504 (M+1). ¹H NMR (400 MHz, CDCl₃) δ8.09 (d, J=8.2 Hz, 1H), 7.43 (s, 1H), 7.39 (dd, J=1.4, 8.0 Hz, 1H), 7.15(br d, J=2.0 Hz, 3H), 7.06-7.00 (m, 1H), 4.83 (br s, 1H), 4.23 (br s,1H), 4.07 (s, 2H), 4.03-3.91 (m, 3H), 3.75-3.78 (m, 1H), 3.54 (s, 1H),3.51-3.45 (m, 1H), 3.04-2.91 (m, 2H), 2.88-2.79 (m, 1H), 2.38-2.22 (m,3H), 2.09-1.95 (m, 3H), 1.93-1.86 (m, 1H), 1.78 (m, 1H), 1.39 (s, 6H)

The other isomer was prepared using the methodology herein and thegeneral procedure described above in step 7 starting with tert-butyl(S)-3-((S)-1-hydroxy-2-(6-((1R,3S,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateto afford2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3S,5S)-3-hydroxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(Example 22): MS: 504 (M+1). ¹H NMR (400 MHz, CDCl₃) δ 8.08 (d, J=7.8Hz, 1H), 7.41 (s, 1H), 7.39-7.31 (m, 1H), 7.14 (d, J=2.7 Hz, 3H), 7.03(br d, J=5.1 Hz, 1H), 4.81 (br s, 1H), 4.22 (br s, 1H), 4.09-3.94 (m,3H), 3.88-3.75 (m, 3H), 3.60-3.49 (m, 2H), 2.99-2.73 (m, 3H), 2.37-2.19(m, 3H), 2.06-1.93 (m, 3H), 1.91-1.85 (m, 1H), 1.73-1.79 (m, 1H), 1.37(s, 6H).

Examples 23-24 Example 23:2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3R,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-oneand Example 24:2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3S,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one

Step 1:

To a mixture of2-(2-((S)-2-(tert-butoxycarbonyl-1,2,3,4-tetrahydroisoquinolin-3-yl)-2-hydroxyethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylicacid (100 mg, 0.202 mmol),(1R,3r,5S)-3-methoxy-8-azabicyclo[3.2.1]octane (34.3 mg, 0.243 mmol) andDIEA (0.106 ml, 0.607 mmol) in DMF (2.0 ml) was added2,4,6-tripropyl-1,3,5,2,4,6-trioxatriphosphinane 2,4,6-trioxide (257 mg,0.404 mmol). The mixture was stirred at 15° C. for 1 h. The reactionmixture was poured into water and extracted with EtOAc (20 mL). Theorganic layer was concentrated under reduced pressure. It was purifiedby Prep-TLC (silica gel, ethyl acetate/pet. ether) to give tert-butyl(3S)-3-(1-hydroxy-2-(6-((1R,3S,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateas colorless oil. MS: 618 (M+1). The stereoisomers were separated by SFCto give two isomers: tert-butyl(S)-3-((R)-1-hydroxy-2-(6-((1R,3R,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(first eluting) and tert-butyl(S)-3-((S)-1-hydroxy-2-(6-((1R,3S,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(second eluting) as white solid.

Step 2:

A solution of: tert-butyl(S)-3-((R)-1-hydroxy-2-(6-((1R,3R,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(30 mg, 0.049 mmol) in HCl (10 ml of 4 N in EtOAc) was stirred at 15° C.for 2 h. The reaction was concentrated under reduced pressure. Theresulting residue was purified by HPLC (ACN/water with 10 mM ammoniumbicarbonate modifier) to give2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3R,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(Example23) as white solid. MS: 518 (M+1). ¹H NMR (400 MHz, CDCl₃) δ 8.08 (d,J=7.8 Hz, 1H), 7.44-7.34 (m, 2H), 7.14 (br d, J=2.3 Hz, 3H), 7.07-6.96(m, 1H), 4.78 (br s, 1H), 4.06 (s, 2H), 4.00-3.88 (m, 3H), 3.74-3.77 (m,1H), 3.54 (br d, J=9.8 Hz, 2H), 3.49-3.42 (m, 1H), 3.29 (s, 3H),3.07-2.73 (m, 3H), 2.24-1.76 (m, 8H), 1.38 (s, 6H).

Example 24 was prepared using the methodology herein and the generalprocedure described above in Step 2 starting with tert-butyl(S)-3-((S)-1-hydroxy-2-(6-((1R,3S,S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)ethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylateto afford2-((S)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3S,5S)-3-methoxy-8-azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl-3,4-dihydroisoquinolin-1(2H)-one(Example24): MS: 518 (M+1). ¹H NMR (400 MHz, CDCl₃) δ 8.08 (d, J=7.8 Hz, 1H),7.41 (s, 1H), 7.36 (d, J=7.8 Hz, 1H), 7.15 (d, J=2.7 Hz, 3H), 7.07-6.99(m, 1H), 4.77 (br s, 1H), 4.11-3.73 (m, 6H), 3.64-3.46 (33H), 3.294 (s,3H), 3.03-2.72 (m, 3H), 2.21-1.77 (m, 8H), 1.37 (s, 6H). The compoundsin the following table were prepared using the methodology herein andthe general procedure described in Examples 23 and 24.

Ex. No. Structure Chemical Name [M + H]+ 25

2-((R)-2-hydroxy-2-((S)- 1,2,3,4- tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3S,5S)-3- methoxy-8- azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl- 3,4-dihydroisoquinolin- 1(2H)-one 518 26

2-((S)-2-hydroxy-2-((S)- 1,2,3,4- tetrahydroisoquinolin-3-yl)ethyl)-6-((1R,3R,5S)- 3-methoxy-8- azabicyclo[3.2.1]octane-8-carbonyl)-4,4-dimethyl- 3,4-dihydroisoquinolin- 1(2H)-one 518 27

6-(hexahydro-3,6- epiminofuro[3,2-b]furan- 7-carbonyl)-2-((R)-2-hydroxy-2-((S)-1,2,3,4- tetrahydroisoquinolin-3- yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin- 1(2H)-one 504 28

6-(hexahydro-3,6- epiminofuro[3,2-b]furan- 7-carbonyl)-2-((S)-2-hydroxy-2-((S)-1,2,3,4- tetrahydroisoquinolin-3- yl)ethyl)-4,4-dimethyl-3,4-dihydroisoquinolin- 1(2H)-one 504

Example 292-{(2R)-2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-4,4-dimethyl-6-(3-oxo-2-azabicyclo[2.2.2]octan-2-yl)-3,4-dihydroisoquinolin-1(2H)-one

A 1.5 mL microwave vial was charged with (S)-tert-butyl3-((R)-2-(6-bromo-4,4-dimethyl-1-oxo-3,4-dihydroisoquinolin-2(1H)-yl)-1-hydroxyethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate(50 mg, 0.094 mmol), 2-azabicyclo[2.2.2]octan-3-one (17.73 mg, 0.142mmol), cesium carbonate (77 mg, 0.236 mmol), andchloro[(4,5-bis(diphenylphosphino)-9,9-dimethylxanthene)-2-(2′-amino-1,1′-biphenyl)]palladium(II)(9.19 mg, 9.44 μmol). The vial was flushed with nitrogen. Dioxane (472μl) was added and the reaction mixture was allowed to stir overnight at90° C. The reaction was then filtered through a syringe filter,concentrated and taken up in 2 mL 1:1 DCM/TFA and allowed to stir for 30minutes at rt. The reaction was concentrated in vacuo, dissolved in 1 mLDMSO, and purified by mass guided reverse phase HPLC (ACN/water+0.1% TFAmodifier) to afford2-(2-((R)-2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-4,4-dimethyl-1-oxo-1,2,3,4-tetrahydroisoquinolin-6-yl)-2-azabicyclo[2.2.2]octan-3-one,TFA salt as a clear resin. MS: 474 (M+1). ¹H NMR (500 MHz, DMSO-d₆) δ9.37-9.26 (m, 1H), 9.04-8.92 (m, 1H), 7.87 (m, 1H), 7.58-7.42 (m, 1H),7.37-7.19 (m, 3H), 5.90 (s, 1H), 4.41 (m, 1H), 4.26 (m, 2H), 4.1 (m,1H), 4.1-3.7 (m, 4H), 3.6-3.5 (m, 2H), 3.45 (m, 1H), 3.42-3.30 (m, 1H),3.18 (s, 2H), 1.85-1.73 (m, 6H), 1.38-1.25 (m, 6H).

Example 302-{2-hydroxy-2-[(3S)-1,2,3,4-tetrahydroisoquinolin-3-yl]ethyl}-6-(3-oxa-8-azabicyclo[3.2.1]oct-8-ylcarbonyl)-3,4-dihydroisoquinolin-1(2H)-one

Step 1:

To a solution of 6-bromo-3,4-dihydroisoquinolin-1(2H)-one (650 mg, 2.88mmol) and KOAc (847 mg, 8.63 mmol) in EtOH (10 mL) was added PdCl₂(dppf)(210 mg, 0.288 mmol) under N₂ atmosphere. The mixture was degassed andbackfilled with CO (three times). The resulting mixture was stirredunder CO (50 psi) at 70° C. for 16 h. The mixture was concentrated underreduced pressure. The residue was purified by flash silica gelchromatography (eluting with ethyl acetate/MeOH gradient) to affordethyl 1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylate as a brownsolid. MS: 220 (M+1).

Step 2:

To a solution of ethyl1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylate (600 mg, 2.74 mmol)in MeOH (5 mL) and water (3 mL) was added lithium hydroxide hydrate (149mg, 3.56 mmol). The mixture was stirred at 20° C. for 1.5 h and thenconcentrated under reduced pressure. The resulting residue was dissolvedin EtOAc (6 mL) and water (5 mL). The resulting slurry was filtered andthe filter cake collected to give1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylic acid as brown solidwhich was used in next step without further purification. MS: 192 (M+1).¹H NMR (400 MHz, CD₃OD) δ 7.91-8.02 (m, 3H), 3.51 (t, J=6.7 Hz, 2H),3.03 (t, J=6.6 Hz, 2H).

Step 3:

To a solution of 1-oxo-1,2,3,4-tetrahydroisoquinoline-6-carboxylic acid(200 mg, 1.046 mmol) in DMF (4 mL) was added HATU (597 mg, 1.569 mmol)and DIEA (0.365 mL, 2.092 mmol). The reaction was stirred for 5 mins at20° C. under N₂. 3-oxa-8-azabicyclo[3.2.1]octane hydrochloride (157 mg,1.046 mmol) was added and the mixture was stirred at 20° C. for 3 h. Thereaction was then dissolved in water (20 mL) and EtOAc (25 mL). Theorganic layer was separated and the aqueous was re-extracted with EtOAc(20 mL×3). The combined organic layers were washed with brine (30 mL),dried over anhydrous Na₂SO₄, filtered, and concentrated. The resultingresidue was purified by column chromatography on silica gel (elutingwith ethyl acetate/MeOH gradient) to afford to give6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-3,4-dihydroisoquinolin-1(2H)-oneas brown oil. MS: 287 (M+1).

Step 4:

To a solution of NaH (52.4 mg, 1.31 mmol) in DMF (4 mL) was added6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-3,4-dihydroisoquinolin-1(2H)-one(250 mg, 0.873 mmol). The reaction mixture was allowed to stir at 20° C.for 0.5 h. Then (3S)-tert-butyl3-(oxiran-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (240 mg, 0.873mmol) was added to the mixture. The reaction mixture was stirred at 80°C. for 18 h. The residue was purified by HPLC (ACN/water with 10 mMNH₄HCO₃ modifier) to give6-(3-oxa-8-azabicyclo[3.2.1]octane-8-carbonyl)-2-(2-hydroxy-2-((S)-1,2,3,4-tetrahydroisoquinolin-3-yl)ethyl)-3,4-dihydroisoquinolin-1(2H)-oneas brown oil. MS: 462 (M+1). H NMR (400 MHz, CD₃D) δ 8.01 (d, J=8.02 Hz,1H), 7.45 (d, J=8.02 Hz, 1H), 7.40 (s, 1H), 7.07-7.14 (m, 3H), 7.01-7.06(m, 1H), 4.63 (br s, 1H), 4.01-4.09 (m, 2H), 3.97-4.01 (m, 1H),3.90-3.97 (m, 2H), 3.74-3.88 (m, 4H), 3.63-3.73 (m, 2H), 3.50-3.63 (m,2H), 3.04-3.14 (m, 2H), 2.77-2.95 (m, 3H), 1.92-2.10 (m, 4H).

PRMT5-MEP50 Enzyme Methylation Assay

PRMT5/MEP50 biochemical assay is a direct measurement of the methylationactivity of the enzyme complex on a short peptide substrate derived fromthe N-terminus of H4 histone. The methylation experiment is performedwith recombinant protein. The assessment of inhibitory effect (IC₅₀) ofsmall molecules is measured by the effectiveness of the compounds toinhibit this reaction.

In this assay, the potency (IC₅₀) of each compound was determined from atwenty-point (1:2 serial dilution; top compound concentration of 100000nM) titration curve using the following outlined procedure. To each wellof a white ProxiPlus 384 well-plate, 100 nL of compound (1% DMSO infinal assay volume of 10 μL) was dispensed, followed by the addition of8 μL of 1× assay buffer (50 mM Bicine pH 8.0, 1 mM DTT, 0.004% Tween20,0.01% BSA) containing 0.5 nM of Full-length (FL)-PRMT5-MEP50 enzymecomplex (recombinant proteins from baculovirus-transfected Sf21 cells:FL-PRMT5; MW=73837 kDa and FL-MEP50; MW=38614). Plates were sealed andplaced in a 37° C. humidified chamber for 30 minutes pre-incubation withcompounds. Subsequently, each reaction was initiated by the addition of2 μL 1× assay buffer containing 75 nM biotinylated H4R3(Mel) peptide,and 15 μM S-(5′-Adenosyl)-L-Methionine Chloride (SAM). The finalreaction in each well of 10 μL consists of 0.5 nM PRMT5-MEP50, 75 nMbiotinylated-peptide, and 15 μM SAM. Methylation reactions were allowedto proceed for 150 minutes in a sealed plate at 37° C. Reactions wereimmediately quenched by the addition of 1 μL of 10% formic acid. Plateswere then frozen and shipped to SAMDI™ Tech Inc. to determine thepercent conversion from K4R3(Mel) to K4R3(Me2). IC₅₀ values weredetermined by 7 parameters biphasic fit model plotting the percentproduct conversion vs. (Log₁₀) compound concentrations.

PRMT5 Cell Target Engagement (TE) Assay

The PRMT5 TE assay is a biomarker assay for identifying compounds thatinhibit symmetric dimethylation of arginine (SDMA) of PRMT5 substrates.This assay detects symmetrically dimethylated nuclear proteins usinghigh content imaging technology. Detection of the expression ofsymmetrically dimethylated nucleo proteins is through a mixture ofprimary rabbit monoclonal antibodies to SDMA (CST 13222), which in turnrecognized by an Alexafluor 488 dye-conjugated anti-rabbit IgG secondaryantibody. The IN Cell Analyzer 2200 measures nuclear Alexafluor 488fluorescent dye intensity that is directly related to the level ofexpression of symmetrically dimethylated nuclear proteins at the singlecell level. Nuclear AF488 dye intensities are compared to the mean valuefor DMSO treated cells (MIN) to report percent of inhibition for eachcompound-treated well.

In this assay, the cell potency (EC₅₀) of each compound was determinedfrom a ten point (1:3 serial dilution; top compound concentration of10000 nM) titration curve using the following outlined procedure. Eachwell of a BD falcon collagen coated black/clear bottom 384-well platewas seeded with 4000 MCF-7 cells and allowed to attach for 5 hours.Media from cell plate was removed at 0.5 mm above the bottom of theplate and replaced with 30 μL of fresh media containing 1.2× compoundsin 0.1% DMSO. Cells were treated for 3 days in 37° C. CO₂ incubator. Onday 3, cells were fixed with Cytofix, permeablized with 0.4%Triton-X-100/Cytofix, and washed with D-PBS without Ca/Mg. Cells wereblocked with Licor Odessey blocking reagent for one hour at roomtemperature, followed by incubation with anti-SDMA (1:1000) antibody at4° C. overnight. 1 antibody was removed, followed by three washings withDPBS without Ca/Mg and 0.05% Tween20. Hoechst (5 μg/ml), Cell Mask deepstain (1:2000) and Alexa488-conjugated goat anti-rabbit IgG (2 μg/mL)was added for 1 hour at room temperature. A final washing step (threewashes) was performed before sealing plate for imaging on In CellAnalyzer 2200. Images from analyzer were uploaded to Columbus (at WP)for image analysis. IC₅₀ values were determined by 4 parameters robustfit of percent fluorescence units vs. (Log₁₀) compound concentrations.

Enzyme TE Ex. Methylation Assay Assay No. (IC₅₀_1, nM; IC₅₀_2, nM)(EC₅₀, nM) 1 5.38; 3741 2856 2 0.21; 171.7 212.7 3 411.1; 21280 10000 445.8; 25790 10000 5 465.4; 100000 10000 6 3.82; 5077 1445 7 0.31; 313.4143.1 8 48.96; 28460 7682 9 0.47; 421.7 29.49 10 0.40; 373.6 22.63 110.29; 127.4 17.0 12 0.63; 253.3 29.79 13 0.36; 270.1 29.97 14 0.24;31.67 2.52 15 9.54; 3208 120.4 16 17.64; 8835 829.2 17 0.61; 368.2 199.818 225.1; 100000 10000 19 0.53; 137.6 9.5 20 12.93; 4067 153.3 21 0.52;88.1 5.52 22 50.12; 15310 5279 23 0.50; 97.72 2.14 24 11.31; 7499 264325 0.44; 50.12 3.81 26 24.27; 14620 4631 27 0.35; 114.8 18.64 28 34.28;34280 10000 29 0.84; 358.9 74.36 30 16.57; 6276 868.7

What is claimed is:
 1. (canceled)
 2. (canceled)
 3. (canceled) 4.(canceled)
 5. (canceled)
 6. (canceled)
 7. A compound selected from oneof the following,

or a pharmaceutically acceptable salt thereof.
 8. A composition fortreating cancer comprising a compound of claim 7, or a pharmaceuticallyacceptable salt thereof, and a pharmaceutically acceptable carrier.
 9. Amethod for treating cancer comprising administering to a patient in needthereof a composition of claim
 8. 10. (canceled)